Soybean Cultivar 2163154

ABSTRACT

A soybean cultivar designated 2163154 is disclosed. Embodiments include the seeds of soybean 2163154, the plants of soybean 2163154, to plant parts of soybean 2163154, and methods for producing a soybean plant produced by crossing soybean 2163154 with itself or with another soybean variety. Embodiments include methods for producing a soybean plant containing in its genetic material one or more genes or transgenes and the transgenic soybean plants and plant parts produced by those methods. Embodiments also relate to soybean cultivars, breeding cultivars, plant parts, and cells derived from soybean 2163154, methods for producing other soybean cultivars, lines or plant parts derived from soybean 2163154, and the soybean plants, varieties, and their parts derived from use of those methods. Embodiments further include hybrid soybean seeds, plants, and plant parts produced by crossing 2163154 with another soybean cultivar.

BACKGROUND

All publications cited in this application are herein incorporated byreference.

There are numerous steps in the development of any novel, desirableplant germplasm. Plant breeding begins with the analysis and definitionof problems and weaknesses of the current germplasm, the establishmentof program goals, and the definition of specific breeding objectives.The next step is selection of germplasm that possesses the traits tomeet the program goals. The goal is to combine in a single variety animproved combination of desirable traits from the parental germplasm.These important traits may include higher seed yield, resistance todiseases and insects, better stems and roots, tolerance to drought andheat, and better agronomic quality.

Soybean, Glycine max (L.) Merr., is an important and valuable fieldcrop. Thus, a continuing goal of soybean plant breeders is to developstable, high yielding soybean cultivars that are agronomically sound.The reasons for this goal are to maximize the amount of grain producedon the land used and to supply food for both animals and humans. Toaccomplish this goal, the soybean breeder must select and developsoybean plants that have traits that result in superior cultivars.

The foregoing examples of the related art and limitations relatedtherewith are intended to be illustrative and not exclusive. Otherlimitations of the related art will become apparent to those of skill inthe art upon a reading of the specification.

SUMMARY

It is to be understood that the embodiments include a variety ofdifferent versions or embodiments, and this Summary is not meant to belimiting or all-inclusive. This Summary provides some generaldescriptions of some of the embodiments, but may also include some morespecific descriptions of other embodiments.

An embodiment provides a soybean cultivar designated 2163154. Anotherembodiment relates to the seeds of soybean cultivar 2163154, to theplants of soybean cultivar 2163154 and to methods for producing asoybean plant produced by crossing soybean cultivar 2163154 with itselfor another soybean cultivar, and the creation of variants by mutagenesisor transformation of soybean cultivar 2163154.

Any such methods using the soybean cultivar 2163154 are a furtherembodiment: selfing, backcrosses, hybrid production, crosses topopulations, and the like. All plants produced using soybean cultivar2163154 as at least one parent are within the scope of the embodiments.Advantageously, soybean cultivar 2163154 could be used in crosses withother, different soybean plants to produce first generation (F₁) soybeanhybrid seeds and plants with superior characteristics.

Another embodiment provides for single or multiple gene converted plantsof soybean cultivar 2163154. The transferred gene(s) may be a dominantor recessive allele. The transferred gene(s) may confer such traits asherbicide resistance, insect resistance, resistance for bacterial,fungal, or viral disease, male fertility, male sterility, enhancednutritional quality, modified fatty acid metabolism, modifiedcarbohydrate metabolism, modified seed yield, modified oil percent,modified protein percent, modified lodging resistance, modifiedshattering, modified iron-deficiency chlorosis, and industrial usage.The gene may be a naturally occurring soybean gene or a transgeneintroduced through genetic engineering techniques.

Another embodiment provides for regenerable cells for use in tissueculture of soybean cultivar 2163154. The tissue culture may be capableof regenerating plants having all the physiological and morphologicalcharacteristics of the foregoing soybean plant, and of regeneratingplants having substantially the same genotype as the foregoing soybeanplant. The regenerable cells in such tissue cultures may be embryos,protoplasts, meristematic cells, callus, pollen, leaves, ovules,anthers, cotyledons, hypocotyl, pistils, roots, root tips, flowers,seeds, petiole, pods, or stems. Still a further embodiment provides forsoybean plants regenerated from the tissue cultures of soybean cultivar2163154.

Another embodiment provides for a method of editing the genome ofsoybean cultivar plant 2163154, said method comprising editing thegenome of the plant, or plant part thereof, of soybean cultivar 2163154,wherein said method is selected from the group comprising zinc fingernucleases, transcription activator-like effector nucleases (TALENs),engineered homing endonucleases/meganucleases, and the clusteredregularly interspaced short palindromic repeat (CRISPR)-associatedprotein9 (Cas9) system.

The soybean seed of soybean cultivar 2163154 may be provided as anessentially homogeneous population of soybean cultivar 2163154.Essentially homogeneous populations of seed are generally free fromsubstantial numbers of other seed.

As used herein, “at least one,” “one or more,” and “and/or” areopen-ended expressions that are both conjunctive and disjunctive inoperation. For example, each of the expressions “at least one of A, Band C,” “at least one of A, B, or C,” “one or more of A, B, and C,” “oneor more of A, B, or C” and “A, B, and/or C” means A alone, B alone, Calone, A and B together, A and C together, B and C together, or A, B andC together.

As used herein, “sometime” means at some indefinite or indeterminatepoint of time. So for example, as used herein, “sometime after” meansfollowing, whether immediately following or at some indefinite orindeterminate point of time following the prior act.

Various embodiments are set forth in the Detailed Description asprovided herein and as embodied by the claims. It should be understood,however, that this Summary does not contain all of the aspects andembodiments, is not meant to be limiting or restrictive in any manner,and that embodiment(s) as disclosed herein is/are understood by those ofordinary skill in the art to encompass obvious improvements andmodifications thereto.

In addition to the exemplary aspects and embodiments described above,further aspects and embodiments will become apparent by study of thefollowing descriptions.

DEFINITIONS

In the description and tables herein, a number of terms are used. Inorder to provide a clear and consistent understanding of thespecification and claims, including the scope to be given such terms,the following definitions are provided:

Cotyledon. A cotyledon is a type of seed leaf. The cotyledon containsthe food storage tissues of the seed.

Embryo. The embryo is the small plant contained within a mature seed.

F₃. The “F₃” symbol denotes a generation resulting from the selfing ofthe F₂ generation along with selection for type and rogueing ofoff-types. The “F” number is a term commonly used in genetics, anddesignates the number of the filial generation. The “F₃” generationdenotes the offspring resulting from the selfing or self mating ofmembers of the generation having the next lower “F” number, that is, the“F₂” generation.

Gene. Gene refers to a segment of nucleic acid. A gene can be introducedinto a genome of a species, whether from a different species or from thesame species, using transformation, gene editing techniques, or variousbreeding methods.

Hilum. Hilum refers to the scar left on the seed that marks the placewhere the seed was attached to the pod prior to the seed beingharvested.

Hypocotyl. A hypocotyl is the portion of an embryo or seedling betweenthe cotyledons and the root. Therefore, it can be considered atransition zone between shoot and root.

Iron Deficiency Chlorosis. Iron deficiency chlorosis (IDC) is ayellowing of the leaves caused by a lack of iron in the soybean plant.Iron is essential in the formation of chlorophyll, which gives plantstheir green color. In high pH soils iron becomes insoluble and cannot beabsorbed by plant roots. Soybean cultivars differ in their geneticability to utilize the available iron. A score of 9 means no stunting ofthe plants or yellowing of the leaves and a score of 1 indicates theplants are dead or dying caused by iron deficiency, a score of 5 meansplants have intermediate health with some leaf yellowing.

Linoleic Acid Percent. Linoleic acid is one of the five most abundantfatty acids in soybean seeds. It is measured by gas chromatography andis reported as a percent of the total oil content.

Locus. A locus confers one or more traits such as, for example, malesterility, herbicide tolerance, insect resistance, disease resistance,waxy starch, modified fatty acid metabolism, modified phytic acidmetabolism, modified carbohydrate metabolism, and modified proteinmetabolism. The trait may be, for example, conferred by a naturallyoccurring gene introduced into the genome of the variety bybackcrossing, a natural or induced mutation, or a transgene introducedthrough genetic transformation techniques. A locus may comprise one ormore alleles integrated at a single chromosomal location.

Lodging Resistance. Lodging resistance refers to the relative presenceof the plant lying on or toward the ground and is on a 1 to 5 scoringbasis. A lodging score of 5 would indicate the plant is basically lyingon the ground. A score of 1 indicates that most or all the plants in arow are standing prostrate.

Maturity Date. Plants are considered mature when 95% of the pods havereached their mature color. The number of days are calculated eitherfrom August 31 or from the planting date.

Maturity Group. Maturity group refers to an agreed-on industry divisionof groups of varieties based on zones in which they are adapted,primarily according to day length or latitude. They consist of very longday length varieties (Groups 000, 00, 0), and extend to very short-daylength varieties (Groups VII, VIII, IX, X).

Oil or Oil Percent. Soybean seeds contain a considerable amount of oil.Oil is measured by NIR spectrophotometry and is reported as a percentagebasis.

Oleic Acid Percent. Oleic acid is one of the five most abundant fattyacids in soybean seeds and is measured by gas chromatography and isreported as a percent of the total oil content.

Palmitic Acid Percent. Palmitic acid is one of the five most abundantfatty acids in soybean seeds and is measured by gas chromatography andis reported as a percent of the total oil content.

Phytophthora Tolerance. Tolerance to Phytophthora root rot is rated on ascale of 1 to 9, with a score of 9 being the best or highest toleranceranging down to a score of 1 which indicates the plants have notolerance to Phytophthora.

Plant. Plant includes reference to an immature or mature whole plant,including a plant from which seed, grain, or anthers have been removed.Seed or embryo that will produce the plant is also considered to be theplant.

Plant Height. Plant height is taken from the top of the soil to the topnode of the plant and is measured in inches or centimeters.

Plant Parts. Plant parts (or a soybean plant, or a part thereof)includes but is not limited to protoplasts, cells, leaves, stems, roots,root tips, anthers, pistils, seed, grain, embryo, pollen, ovules,cotyledon, hypocotyl, pod, flower, shoot, tissue, petiole, cells,meristematic cells, and the like.

Pod. Pod refers to the fruit of a soybean plant. It consists of the hullor shell (pericarp) and the soybean seeds.

Progeny. Progeny includes an F₁ soybean plant produced from the cross oftwo soybean plants where at least one plant includes soybean cultivar2163154 and progeny further includes, but is not limited to, subsequentF₂, F₃, F₄, F₅, F₆, F₇, F₈, F₉, and F₁₀ generational crosses with therecurrent parental line.

Protein Percent. Soybean seeds contain a considerable amount of protein.Protein is generally measured by NIR spectrophotometry and is reportedon an as is percentage basis.

Pubescence. Pubescence refers to a covering of very fine hairs closelyarranged on the leaves, stems, and pods of the soybean plant.

Quantitative Trait Loci (QTL). Quantitative trait loci (QTL) refer togenetic loci that control to some degree numerically representabletraits that are usually continuously distributed.

Relative Maturity (RM). The term relative maturity is a numerical valuethat is assigned to a soybean variety based on comparisons with thematurity values of other varieties. The number preceding the decimalpoint in the RM refers to the maturity group. The number following thedecimal point refers to the relative earliness or lateness within eachmaturity group. For example, a 3.0 is an early group III variety, whilea 3.9 is a late group III variety.

Seed Yield (Bushels/Acre). The yield in bushels/acre is the actual yieldof the grain at harvest.

Seeds Per Pound. Soybean seeds vary in seed size; therefore, the numberof seeds required to make up one pound also varies. The number of seedsper pound affects the pounds of seed required to plant a given area andcan also impact end uses.

Single Gene Converted (Conversion). Single gene converted (conversion)plants refers to plants which are developed by a plant breedingtechnique called backcrossing wherein essentially all of the desiredmorphological and physiological characteristics of a variety arerecovered in addition to the single gene transferred into the varietyvia the backcrossing technique or via genetic engineering. Byessentially all of the morphological and physiological characteristics,it is meant that the characteristics of a plant are recovered that areotherwise present when compared in the same environment, other than anoccasional variant trait that might arise during backcrossing or directintroduction of a transgene.

Sulfonylurea Reaction. Sulfonylurea reaction refers to a plant'stolerance, resistance or susceptibility to sulfonylurea herbicides andrefers to a plant which contains the ALS gene, which confers resistanceto some of the sulfonylurea herbicides.

Trypsin. Trypsin is a digestive enzyme, specifically, a pancreaticserine protease enzyme with substrate specificity based upon positivelycharged lysine and arginine side chains and is excreted by the pancreas.Trypsin aids in the digestion of food proteins and other biologicalprocesses.

Trypsin inhibitor units. Trypsin inhibitor units or abbreviated as TIU,is an assay measuring the quantity of trypsin inhibitor in a soybeanseed or soybean product thereof. Measurement of trypsin inhibitor unitsis a technique well-known in the art.

DETAILED DESCRIPTION

Soybean cultivar 2163154 is an early-group III maturity variety.Additionally, soybean cultivar 2163154 is resistant to Soybean CystNematode Race 3, tolerant to sulfonylurea (ALS inhibitor), and issusceptible to Phytophthora root rot.

Some of the selection criteria used for various generations include:seed yield, lodging resistance, emergence, disease tolerance, maturity,late season plant intactness, plant height, and shattering resistance.

Soybean cultivar 2163154 has shown uniformity and stability, asdescribed in the following variety description information. Soybeancultivar 2163154 has been self-pollinated a sufficient number ofgenerations with careful attention to uniformity of plant type and hasbeen increased with continued observation for uniformity.

Soybean cultivar 2163154 has the following morphologic and othercharacteristics based primarily on data collected at the followinglocations: Newburg, Iowa; Atlantic, Iowa; Ralston, Iowa; Washington,Iowa; Walcott, Iowa; Winterset, Iowa; Wahpeton, N. Dak.; Chestnut, Ill.;Monmouth, Ill.; Shipman, Ill.; Vandalia, Ill.; Albion, Nebr.; Beatrice,Nebr.; Stromsburg, Nebr.; Napoleon, Ohio; Lenox, Iowa; Minburn, Iowa;Gilman, Ill.; Crawfordsville, Iowa; Essex, Iowa; Mount Pleasant, Iowa;Ottumwa, Iowa; Pleasantville, Iowa; Farmersville, Ill.; Mattoon, Ill.;Eureka, Ill.; Ivesdale, Ill.; Queenstown, Md.; Rock Hall, Md.; Gallatin,Mo.; Ellsworth, Iowa; Bloomington, Ill.; Champaign, Ill.; Geneseo, Ill.;Mendota, Ill.; Bluffton, Ind.; Greensburg, Ind.; Marion, Ind.; Wabash,Ind.; Norfolk, Nebr.; Archbold, Ohio; Covington, Ohio; Delphos, Ohio;Greenville, Ohio; Earlham, Iowa; Shell Rock, Iowa; Humboldt, Iowa; NoraSprings, Iowa.

TABLE 1 VARIETY DESCRIPTION INFORMATION Hypocotyl Color: Clear Seed CoatColor (Mature Seed): Clear Seed Coat Luster (Mature Hand Shelled Seed):Dull Seed Color (Mature Seed): Yellow Leaflet Shape: Ovate Growth Habit:Indeterminate Flower Color: Purple Hilum Color (Mature Seed): ImperfectYellow Plant Pubescence Color: Tawny Pod Wall Color: Brown MaturityGroup: 3 Relative Maturity: 3.1 Plant Lodging Score: 2.3 Plant Height(cm): 96 Percent Protein: 42.7% dry weight Percent Oil: 20.4% dry weightPhysiological Responses (known resistances/susceptibility): Resistant toSoybean Cyst Nematode (Race 3), tolerant to sulfonylurea (ALSinhibitor), and is susceptible to Phytophthora root rot

In Table 2, the yield of soybean cultivar 2163154 is compared with theyield of soybean cultivars P93Y21, Becks 327, and e3494 from 2015 to2018 in the United States in side-by-side trials. Column one shows thesoybean cultivar designations, column two shows the year, column threeshows the number of locations, column four shows the number ofobservations, and column five shows the yield in bushels per acre.

TABLE 2 Yield comparison with commercial cultivars Soybean Cultivar Year# of Locations # of Observations Yield 2163154 2016 to 2016 16 16 61.9P93Y21 2015 to 2016 16 16 60.3 2163154 2016 and 2018 20 20 61.8 Becks327 2016 and 2018 20 20 57.9 2163154 2015 to 2018 82 82 58.8 e3494 2015to 2018 82 82 61.3

In Table 3, the characteristics of soybean cultivar 2163154 are comparedwith soybean cultivars P93Y21, Becks 327, and e3494.

TABLE 3 Comparison of characteristics with commercial cultivars SoybeanCultivar Characteristic 2163154 P93Y21 Becks 327 e3494 Flower colorPurple White Purple White Pubescence color Tawny Light tawny Grey Lighttawny Hilum color Imperfect Black Yellow Black yellow Pod wall colorBrown Brown Brown Brown Soybean Cyst Resistant Resistant Nematodereaction Phytophthora Susceptible Susceptible reaction Sulfonylureareaction Tolerant Not tolerant

Breeding With Soybean Cultivar 2163154

The complexity of inheritance influences choice of the breeding method.Backcross breeding is used to transfer one or a few favorable genes fora highly heritable trait into a desirable cultivar. This approach hasbeen used extensively for breeding disease-resistant cultivars. Variousrecurrent selection techniques are used to improve quantitativelyinherited traits controlled by numerous genes. The use of recurrentselection in self-pollinating crops depends on the ease of pollination,the frequency of successful hybrids from each pollination, and thenumber of hybrid offspring from each successful cross.

Promising advanced breeding lines are thoroughly tested and compared toappropriate standards in environments representative of the commercialtarget area(s) for three or more years. The best lines are candidatesfor new commercial cultivars; those still deficient in a few traits maybe used as parents to produce new populations for further selection.

These processes, which lead to the final step of marketing anddistribution, usually take from eight to twelve years from the time thefirst cross is made. Therefore, development of new cultivars is atime-consuming process that requires precise forward planning, efficientuse of resources, and a minimum of changes in direction.

A most difficult task is the identification of individuals that aregenetically superior, because for most traits the true genotypic valueis masked by other confounding plant traits or environmental factors.One method of identifying a superior plant is to observe its performancerelative to other experimental plants and to a widely grown standardcultivar. If a single observation is inconclusive, replicatedobservations provide a better estimate of its genetic worth.

The goal of soybean plant breeding is to develop new and superiorsoybean cultivars and hybrids. The breeder initially selects and crossestwo or more parental lines, followed by repeated selfing and selection,producing many new genetic combinations. The breeder can theoreticallygenerate billions of different genetic combinations via crossing,selection, selfing and mutations. Therefore, a breeder will neverdevelop the same line, or even very similar lines, having the samesoybean traits from the exact same parents.

Each year, the plant breeder selects the germplasm to advance to thenext generation. This germplasm is grown under different geographical,climatic and soil conditions and further selections are then made duringand at the end of the growing season. The cultivars that are developedare unpredictable because the breeder's selection occurs in environmentswith no control at the DNA level, and with millions of differentpossible genetic combinations being generated. A breeder of ordinaryskill in the art cannot predict the final resulting lines he develops,except possibly in a very gross and general fashion. The same breedercannot produce the same cultivar twice by using the same originalparents and the same selection techniques. This unpredictability resultsin the expenditure of large amounts of research monies to developsuperior new soybean cultivars.

The development of new soybean cultivars requires the development andselection of soybean varieties, the crossing of these varieties andselection of superior hybrid crosses. The hybrid seed is produced bymanual crosses between selected male-fertile parents or by using malesterility systems. These hybrids are selected for certain single genetraits such as pod color, flower color, pubescence color or herbicideresistance which indicate that the seed is truly a hybrid. Additionaldata on parental lines, as well as the phenotype of the hybrid,influence the breeder's decision whether to continue with the specifichybrid cross.

Breeding programs combine desirable traits from two or more cultivars orvarious broad-based sources into breeding pools from which cultivars aredeveloped by selfing and selection of desired phenotypes. Pedigreebreeding is used commonly for the improvement of self-pollinating crops.Two parents that possess favorable, complementary traits are crossed toproduce an F₁. An F₂ population is produced by selfing one or severalF₁s. Selection of the best individuals may begin in the F₂ population;then, beginning in the F₃, the best individuals in the best families areselected. Replicated testing of families can begin in the F₄ generationto improve the effectiveness of selection for traits with lowheritability. At an advanced stage of inbreeding (i.e., F₆ and F₇), thebest lines or mixtures of phenotypically similar lines are tested forpotential release as new cultivars.

Using Soybean Cultivar 2163154 to Develop other Soybean Varieties

Soybean varieties such as soybean cultivar 2163154 are typicallydeveloped for use in seed and grain production. However, soybeanvarieties such as soybean cultivar 2163154 also provide a source ofbreeding material that may be used to develop new soybean varieties.Plant breeding techniques known in the art and used in a soybean plantbreeding program include, but are not limited to, recurrent selection,mass selection, bulk selection, mass selection, backcrossing, pedigreebreeding, open pollination breeding, restriction fragment lengthpolymorphism enhanced selection, genetic marker enhanced selection,making double haploids, and transformation. Often combinations of thesetechniques are used. The development of soybean varieties in a plantbreeding program requires, in general, the development and evaluation ofhomozygous varieties. There are many analytical methods available toevaluate a new variety. The oldest and most traditional method ofanalysis is the observation of phenotypic traits, but genotypic analysismay also be used.

Additional Breeding Methods

One embodiment is directed to methods for producing a soybean plant bycrossing a first parent soybean plant with a second parent soybeanplant, wherein the first or second soybean plant is the soybean plantfrom soybean cultivar 2163154. Further, both first and second parentsoybean plants may be from soybean cultivar 2163154. Therefore, anymethods using soybean cultivar 2163154 are part of the embodiments:selfing, backcrosses, hybrid breeding, and crosses to populations. Anyplants produced using soybean cultivar 2163154 as at least one parentare also within the scope of the embodiments. Any such methods usingsoybean variety 2163154 are part of the embodiments: selfing, sibbing,backcrosses, mass selection, pedigree breeding, bulk selection, hybridproduction, crosses to populations, and the like. These methods are wellknown in the art and some of the more commonly used breeding methods aredescribed herein. Descriptions of breeding methods can be found in oneof several reference books (e.g., Allard, Principles of Plant Breeding(1960); Simmonds, Principles of Crop Improvement (1979); Sneep, et al.(1979); Fehr, “Breeding Methods for Cultivar Development,” Chapter 7,Soybean Improvement, Production and Uses, 2^(nd) ed., Wilcox editor(1987)).

The following describes breeding methods that may be used with soybeancultivar 2163154 in the development of further soybean plants. One suchembodiment is a method for developing a cultivar 2163154 progeny soybeanplant in a soybean plant breeding program comprising: obtaining thesoybean plant, or a part thereof, of cultivar 2163154, utilizing saidplant, or plant part, as a source of breeding material, and selecting asoybean cultivar 2163154 progeny plant with molecular markers in commonwith cultivar 2163154 and/or with morphological and/or physiologicalcharacteristics selected from the characteristics listed in Tables 1and/or 2 and/or 3 and/or 4. Breeding steps that may be used in thesoybean plant breeding program include pedigree breeding, backcrossing,mutation breeding, and recurrent selection. In conjunction with thesesteps, techniques such as RFLP-enhanced selection, genetic markerenhanced selection (for example, SSR markers), and the making of doublehaploids may be utilized.

Another method involves producing a population of soybean cultivar2163154 progeny soybean plants, comprising crossing cultivar 2163154with another soybean plant, thereby producing a population of soybeanplants which, on average, derive 50% of their alleles from soybeancultivar 2163154. A plant of this population may be selected andrepeatedly selfed or sibbed with a soybean cultivar resulting from thesesuccessive filial generations. One embodiment is the soybean cultivarproduced by this method and that has obtained at least 50% of itsalleles from soybean cultivar 2163154.

One of ordinary skill in the art of plant breeding would know how toevaluate the traits of two plant varieties to determine if there is nosignificant difference between the two traits expressed by thosevarieties. For example, see, Fehr and Walt, Principles of CultivarDevelopment, pp. 261-286 (1987). Thus, embodiments include soybeancultivar 2163154 progeny soybean plants comprising a combination of atleast two cultivar 2163154 traits selected from the group consisting ofthose listed in Tables 1 and/or 2 and/or 3 and/or 4 or soybean cultivar2163154 combination of traits listed in the Summary, so that saidprogeny soybean plant is not significantly different for said traitsthan soybean cultivar 2163154 as determined at the 5% significance levelwhen grown in the same environmental conditions. Using techniquesdescribed herein, molecular markers may be used to identify said progenyplant as a soybean cultivar 2163154 progeny plant. Mean trait values maybe used to determine whether trait differences are significant, andpreferably the traits are measured on plants grown under the sameenvironmental conditions. Once such a variety is developed, its value issubstantial since it is important to advance the germplasm base as awhole in order to maintain or improve traits such as yield, diseaseresistance, pest resistance, and plant performance in extremeenvironmental conditions.

Progeny of soybean cultivar 2163154 may also be characterized throughtheir filial relationship with soybean cultivar 2163154, as for example,being within a certain number of breeding crosses of soybean cultivar2163154. A breeding cross is a cross made to introduce new genetics intothe progeny, and is distinguished from a cross, such as a self or a sibcross, made to select among existing genetic alleles. The lower thenumber of breeding crosses in the pedigree, the closer the relationshipbetween soybean cultivar 2163154 and its progeny. For example, progenyproduced by the methods described herein may be within 1, 2, 3, 4, or 5breeding crosses of soybean cultivar 2163154.

As used herein, the term “plant” includes plant cells, plantprotoplasts, plant cell tissue cultures from which soybean plants can beregenerated, plant calli, plant clumps, and plant cells that are intactin plants or parts of plants, such as embryos, pollen, ovules, flowers,pods, leaves, roots, root tips, anthers, cotyledons, hypocotyls,meristematic cells, stems, pistils, petiole, and the like.

Pedigree Breeding

Pedigree breeding starts with the crossing of two genotypes, such assoybean cultivar 2163154 and another soybean variety having one or moredesirable characteristics that is lacking or which complements soybeancultivar 2163154. If the two original parents do not provide all thedesired characteristics, other sources can be included in the breedingpopulation. In the pedigree method, superior plants are selfed andselected in successive filial generations. In the succeeding filialgenerations, the heterozygous condition gives way to homogeneousvarieties as a result of self-pollination and selection. Typically, inthe pedigree method of breeding, five or more successive filialgenerations of selfing and selection is practiced: F₁ to F₂; F₂ to F₃;F₃ to F₄; F₄ to F₅; etc. After a sufficient amount of inbreeding,successive filial generations will serve to increase seed of thedeveloped variety. Preferably, the developed variety compriseshomozygous alleles at about 95% or more of its loci.

Backcross Breeding

Backcross breeding has been used to transfer genes for a simplyinherited, highly heritable trait into a desirable homozygous cultivaror inbred line which is the recurrent parent. The source of the trait tobe transferred is called the donor parent. After the initial cross,individuals possessing the phenotype of the donor parent are selectedand repeatedly crossed (backcrossed) to the recurrent parent. Theresulting plant is expected to have the attributes of the recurrentparent (e.g., cultivar) and the desirable trait transferred from thedonor parent.

In addition to being used to create a backcross conversion, backcrossingcan also be used in combination with pedigree breeding. As discussedpreviously, backcrossing can be used to transfer one or morespecifically desirable traits from one variety, the donor parent, to adeveloped variety called the recurrent parent, which has overall goodagronomic characteristics yet lacks that desirable trait or traits.However, the same procedure can be used to move the progeny toward thegenotype of the recurrent parent, but at the same time retain manycomponents of the nonrecurrent parent by stopping the backcrossing at anearly stage and proceeding with selfing and selection. For example, asoybean variety may be crossed with another variety to produce afirst-generation progeny plant. The first-generation progeny plant maythen be backcrossed to one of its parent varieties to create a BC₁ orBC₂. Progeny are selfed and selected so that the newly developed varietyhas many of the attributes of the recurrent parent and yet several ofthe desired attributes of the nonrecurrent parent. This approachleverages the value and strengths of the recurrent parent for use in newsoybean varieties.

Therefore, an embodiment is a method of making a backcross conversion ofsoybean variety 2163154, comprising the steps of crossing a plant ofsoybean variety 2163154 with a donor plant comprising a desired trait,selecting an F₁ progeny plant comprising the desired trait, andbackcrossing the selected F₁ progeny plant to a plant of soybean variety2163154. This method may further comprise the step of obtaining amolecular marker profile of soybean variety 2163154 and using themolecular marker profile to select for a progeny plant with the desiredtrait and the molecular marker profile of soybean cultivar 2163154. Inone embodiment, the desired trait is a mutant gene, gene, or transgenepresent in the donor parent.

Recurrent Selection and Mass Selection

Recurrent selection is a method used in a plant breeding program toimprove a population of plants. Soybean cultivar 2163154 is suitable foruse in a recurrent selection program. The method entails individualplants cross pollinating with each other to form progeny. The progenyare grown and the superior progeny selected by any number of selectionmethods, which include individual plant, half-sib progeny, full-sibprogeny, and selfed progeny. The selected progeny are cross pollinatedwith each other to form progeny for another population. This populationis planted and again superior plants are selected to cross pollinatewith each other. Recurrent selection is a cyclical process and thereforecan be repeated as many times as desired. The objective of recurrentselection is to improve the traits of a population. The improvedpopulation can then be used as a source of breeding material to obtainnew varieties for commercial or breeding use, including the productionof a synthetic cultivar. A synthetic cultivar is the resultant progenyformed by the intercrossing of several selected varieties.

Mass selection is a useful technique when used in conjunction withmolecular marker enhanced selection. In mass selection, seeds fromindividuals are selected based on phenotype or genotype. These selectedseeds are then bulked and used to grow the next generation. Bulkselection requires growing a population of plants in a bulk plot,allowing the plants to self-pollinate, harvesting the seed in bulk, andthen using a sample of the seed harvested in bulk to plant the nextgeneration. Also, instead of self-pollination, directed pollinationcould be used as part of the breeding program.

Mass and recurrent selections can be used to improve populations ofeither self- or cross-pollinating crops. A genetically variablepopulation of heterozygous individuals is either identified, or created,by intercrossing several different parents. The plants are selectedbased on individual superiority, outstanding progeny, or excellentcombining ability. The selected plants are intercrossed to produce a newpopulation in which further cycles of selection are continued.

Single-Seed Descent

The single-seed descent procedure in the strict sense refers to plantinga segregating population, harvesting a sample of one seed per plant, andusing the one-seed sample to plant the next generation. When thepopulation has been advanced from the F₂ to the desired level ofinbreeding, the plants from which lines are derived will each trace todifferent F₂ individuals. The number of plants in a population declineseach generation due to failure of some seeds to germinate or some plantsto produce at least one seed. As a result, not all of the F₂ plantsoriginally sampled in the population will be represented by a progenywhen generation advance is completed.

Multiple-Seed Procedure

In a multiple-seed procedure, soybean breeders commonly harvest one ormore pods from each plant in a population and thresh them together toform a bulk. Part of the bulk is used to plant the next generation andpart is put in reserve. The procedure has been referred to as modifiedsingle-seed descent or the pod-bulk technique.

The multiple-seed procedure has been used to save labor at harvest. Itis considerably faster to thresh pods with a machine than to remove oneseed from each by hand for the single-seed procedure. The multiple-seedprocedure also makes it possible to plant the same number of seeds of apopulation in each generation of inbreeding. Enough seeds are harvestedto make up for those plants that did not germinate or produce seed.

Mutation Breeding

Mutation breeding is another method of introducing new traits intosoybean variety 2163154. Mutations that occur spontaneously or areartificially induced can be useful sources of variability for a plantbreeder. The goal of artificial mutagenesis is to increase the rate ofmutation for a desired characteristic. Mutation rates can be increasedby many different means including temperature, long-term seed storage,tissue culture conditions, radiation; such as X-rays, Gamma rays (e.g.,cobalt 60 or cesium 137), neutrons, (product of nuclear fission byuranium 235 in an atomic reactor), Beta radiation (emitted fromradioisotopes such as phosphorus 32 or carbon 14), or ultravioletradiation (preferably from 2500 to 2900 nm), or chemical mutagens (suchas base analogues (5-bromo-uracil)), related compounds (8-ethoxycaffeine), antibiotics (streptonigrin), alkylating agents (sulfurmustards, nitrogen mustards, epoxides, ethylenamines, sulfates,sulfonates, sulfones, lactones), azide, hydroxylamine, nitrous acid, oracridines. Once a desired trait is observed through mutagenesis thetrait may then be incorporated into existing germplasm by traditionalbreeding techniques. Details of mutation breeding can be found in Fehr,“Principles of Cultivar Development,” Macmillan Publishing Company(1993). In addition, mutations created in other soybean plants may beused to produce a backcross conversion of soybean cultivar 2163154 thatcomprises such mutation.

Additional methods include, but are not limited to, expression vectorsintroduced into plant tissues using a direct gene transfer method, suchas microprojectile-mediated delivery, DNA injection, electroporation,and the like. More preferably, expression vectors are introduced intoplant tissues by using either microprojectile-mediated delivery with abiolistic device or by using Agrobacterium-mediated transformation.Transformant plants obtained with the protoplasm of the embodiments areintended to be within the scope of the embodiments.

Gene Editing Using CRISPR

Targeted gene editing can be done using CRISPR/Cas9 technology (Saunders& Joung, Nature Biotechnology, 32, 347-355, 2014). CRISPR is a type ofgenome editing system that stands for Clustered Regularly InterspacedShort Palindromic Repeats. This system and CRISPR-associated (Cas) genesenable organisms, such as select bacteria and archaea, to respond to andeliminate invading genetic material. Ishino, Y., et al. J. Bacteriol.169, 5429-5433 (1987). These repeats were known as early as the 1980s inE. coli, but Barrangou and colleagues demonstrated that S. thermophiluscan acquire resistance against a bacteriophage by integrating a fragmentof a genome of an infectious virus into its CRISPR locus. Barrangou, R.,et al. Science 315, 1709-1712 (2007). Many plants have already beenmodified using the CRISPR system, including soybean. See for example,Liu, J., et al. Genome Editing in Soybean with CRISPR/Cas9. Methods MolBiol. 2019. 1917:217-234.

Gene editing can also be done using crRNA-guided surveillance systemsfor gene editing. Additional information about crRNA-guided surveillancecomplex systems for gene editing can be found in the followingdocuments: U.S. Application Publication No. 2010/0076057 (Sontheimer etal., Target DNA Interference with crRNA); U.S. Application PublicationNo. 2014/0179006 (Feng, CRISPR-CAS Component Systems, Methods, andCompositions for Sequence Manipulation); U.S. Application PublicationNo. 2014/0294773 (Brouns et al., Modified Cascade Ribonucleoproteins andUses Thereof); Sorek et al., Annu. Rev. Biochem. 82:273-266, 2013; andWang, S. et al., Plant Cell Rep (2015) 34: 1473-1476.

Therefore, it is another embodiment to use the CRISPR system on soybeancultivar 2163154 to modify traits and resistances or tolerances topests, herbicides, and viruses.

Gene Editing Using TALENs

Transcription activator-like effector nucleases (TALENs) have beensuccessfully used to introduce targeted mutations via repair of doublestranded breaks (DSBs) either through non-homologous end joining (NHEJ),or by homology-directed repair (HDR) and homology-independent repair inthe presence of a donor template. Thus, TALENs are another mechanism fortargeted genome editing using soybean cultivar 2163154. The technique iswell known in the art; see for example Malzahn, Aimee et al. “Plantgenome editing with TALEN and CRISPR” Cell & bioscience vol. 7 21. 24Apr. 2017.

Therefore, it is another embodiment to use the TALENs system on soybeancultivar to modify traits and resistances or tolerances to pests,herbicides, and viruses.

Other Methods of Genome Editing

In addition to CRISPR and TALENs, two other types of engineerednucleases can be used for genome editing: engineered homingendonucleases/meganucleases (EMNs), and zinc finger nucleases (ZFNs).These methods are well known in the art. See for example, Petilino,Joseph F. “Genome editing in plants via designed zinc finger nucleases”In Vitro Cell Dev Biol Plant. 51(1): pp. 1-8 (2015); and Daboussi,Fayza, et al. “Engineering Meganuclease for Precise Plant GenomeModification” in Advances in New Technology for Targeted Modification ofPlant Genomes. Springer Science+Business. pp 21-38 (2015).

Therefore, it is another embodiment to use engineered nucleases onsoybean cultivar to modify traits and resistances or tolerances topests, herbicides, and viruses.

Single-Gene Conversions

When the term “soybean plant” is used in the context of an embodiment,this also includes any single gene conversions of that variety. The termsingle gene converted plant as used herein refers to those soybeanplants which are developed by a plant breeding technique calledbackcrossing wherein essentially all of the desired morphological andphysiological characteristics of a variety are recovered in addition tothe single gene transferred into the variety via the backcrossingtechnique. Backcrossing methods can be used with one embodiment toimprove or introduce a characteristic into the variety. The term“backcrossing” as used herein refers to the repeated crossing of ahybrid progeny back to the recurrent parent, i.e., backcrossing 1, 2, 3,4, 5, 6, 7, 8, or more times to the recurrent parent. The parentalsoybean plant that contributes the gene for the desired characteristicis termed the nonrecurrent or donor parent. This terminology refers tothe fact that the nonrecurrent parent is used one time in the backcrossprotocol and therefore does not recur. The parental soybean plant towhich the gene or genes from the nonrecurrent parent are transferred isknown as the recurrent parent as it is used for several rounds in thebackcrossing protocol (Poehlman & Sleper (1994); Fehr, Principles ofCultivar Development, pp. 261-286 (1987)). In a typical backcrossprotocol, the original variety of interest (recurrent parent) is crossedto a second variety (nonrecurrent parent) that carries the single geneof interest to be transferred. The resulting progeny from this cross arethen crossed again to the recurrent parent and the process is repeateduntil a soybean plant is obtained wherein essentially all of the desiredmorphological and physiological characteristics of the recurrent parentare recovered in the converted plant, in addition to the singletransferred gene from the nonrecurrent parent.

The selection of a suitable recurrent parent is an important step for asuccessful backcrossing procedure. The goal of a backcross protocol isto alter or substitute a single trait or characteristic in the originalvariety. To accomplish this, a single gene of the recurrent variety ismodified or substituted with the desired gene from the nonrecurrentparent, while retaining essentially all of the rest of the desiredgenetic, and therefore the desired physiological and morphologicalconstitution of the original variety. The choice of the particularnonrecurrent parent will depend on the purpose of the backcross; one ofthe major purposes is to add some agronomically important trait to theplant. The exact backcrossing protocol will depend on the characteristicor trait being altered to determine an appropriate testing protocol.Although backcrossing methods are simplified when the characteristicbeing transferred is a dominant allele, a recessive allele may also betransferred. In this instance, it may be necessary to introduce a testof the progeny to determine if the desired characteristic has beensuccessfully transferred.

Many single gene traits have been identified that are not regularlyselected for in the development of a new variety but that can beimproved by backcrossing techniques. Single gene traits may or may notbe transgenic. Examples of these traits include, but are not limited to,male sterility, waxy starch, herbicide resistance, resistance forbacterial, fungal, or viral disease, insect resistance, male fertility,enhanced nutritional quality, industrial usage, yield stability, andyield enhancement. These genes are generally inherited through thenucleus. Several of these single gene traits are described in U.S. Pat.Nos. 5,959,185, 5,973,234, and 5,977,445, the disclosures of which arespecifically hereby incorporated by reference.

Introduction of a New Trait or Locus into Soybean Cultivar 2163154

Variety 2163154 represents a new variety into which a new locus or traitmay be introgressed. Direct transformation and backcrossing representtwo important methods that can be used to accomplish such anintrogression. The term backcross conversion and single locus conversionare used interchangeably to designate the product of a backcrossingprogram.

Backcross Conversions of Soybean Cultivar 2163154

A backcross conversion of soybean cultivar 2163154 occurs when DNAsequences are introduced through backcrossing (Hallauer, et al., “CornBreeding,” Corn and Corn Improvements, No. 18, pp. 463-481 (1988)), withsoybean cultivar 2163154 utilized as the recurrent parent. Bothnaturally occurring and transgenic DNA sequences may be introducedthrough backcrossing techniques. A backcross conversion may produce aplant with a trait or locus conversion in at least two or morebackcrosses, including at least 2 crosses, at least 3 crosses, at least4 crosses, at least 5 crosses, and the like. Molecular marker assistedbreeding or selection may be utilized to reduce the number ofbackcrosses necessary to achieve the backcross conversion. For example,see, Openshaw, S. J., et al., Marker-assisted Selection in BackcrossBreeding, Proceedings Symposium of the Analysis of Molecular Data, CropScience Society of America, Corvallis, Oreg. (August 1994), where it isdemonstrated that a backcross conversion can be made in as few as twobackcrosses.

The complexity of the backcross conversion method depends on the type oftrait being transferred (single genes or closely linked genes ascompared to unlinked genes), the level of expression of the trait, thetype of inheritance (cytoplasmic or nuclear), and the types of parentsincluded in the cross. It is understood by those of ordinary skill inthe art that for single gene traits that are relatively easy toclassify, the backcross method is effective and relatively easy tomanage. (See, Hallauer, et al., Corn and Corn Improvement, Sprague andDudley, Third Ed. (1998)). Desired traits that may be transferredthrough backcross conversion include, but are not limited to, sterility(nuclear and cytoplasmic), fertility restoration, nutritionalenhancements, drought tolerance, nitrogen utilization, altered fattyacid profile, low phytate, industrial enhancements, disease resistance(bacterial, fungal, or viral), insect resistance, and herbicideresistance. In addition, an introgression site itself, such as an FRTsite, Lox site, or other site-specific integration site, may be insertedby backcrossing and utilized for direct insertion of one or more genesof interest into a specific plant variety. In some embodiments, thenumber of loci that may be backcrossed into soybean cultivar 2163154 isat least 1, 2, 3, 4, or 5, and/or no more than 6, 5, 4, 3, or 2. Asingle locus may contain several transgenes, such as a transgene fordisease resistance that, in the same expression vector, also contains atransgene for herbicide resistance. The gene for herbicide resistancemay be used as a selectable marker and/or as a phenotypic trait. Asingle locus conversion of site specific integration system allows forthe integration of multiple genes at the converted loci.

The backcross conversion may result from either the transfer of adominant allele or a recessive allele. Selection of progeny containingthe trait of interest is accomplished by direct selection for a traitassociated with a dominant allele. Transgenes transferred viabackcrossing typically function as a dominant single gene trait and arerelatively easy to classify. Selection of progeny for a trait that istransferred via a recessive allele requires growing and selfing thefirst backcross generation to determine which plants carry the recessivealleles. Recessive traits may require additional progeny testing insuccessive backcross generations to determine the presence of the locusof interest. The last backcross generation is usually selfed to givepure breeding progeny for the gene(s) being transferred, although abackcross conversion with a stably introgressed trait may also bemaintained by further backcrossing to the recurrent parent withselection for the converted trait.

Along with selection for the trait of interest, progeny are selected forthe phenotype of the recurrent parent. The backcross is a form ofinbreeding, and the features of the recurrent parent are automaticallyrecovered after successive backcrosses. Poehlman, Breeding Field Crops,p. 204 (1987). Poehlman suggests from one to four or more backcrosses,but as noted above, the number of backcrosses necessary can be reducedwith the use of molecular markers. Other factors, such as a geneticallysimilar donor parent, may also reduce the number of backcrossesnecessary. As noted by Poehlman, backcrossing is easiest for simplyinherited, dominant, and easily recognized traits.

One process for adding or modifying a trait or locus in soybean variety2163154 comprises crossing soybean cultivar 2163154 plants grown fromsoybean cultivar 2163154 seed with plants of another soybean varietythat comprise the desired trait or locus, selecting F₁ progeny plantsthat comprise the desired trait or locus to produce selected F₁ progenyplants, crossing the selected progeny plants with the soybean cultivar2163154 plants to produce backcross progeny plants, selecting forbackcross progeny plants that have the desired trait or locus and themorphological characteristics of soybean variety 2163154 to produceselected backcross progeny plants, and backcrossing to soybean cultivar2163154 three or more times in succession to produce selected fourth orhigher backcross progeny plants that comprise said trait or locus. Themodified soybean cultivar 2163154 may be further characterized as havingthe physiological and morphological characteristics of soybean variety2163154 listed in Table 1 as determined at the 5% significance levelwhen grown in the same environmental conditions and/or may becharacterized by percent similarity or identity to soybean cultivar2163154 as determined by SSR markers. The above method may be utilizedwith fewer backcrosses in appropriate situations, such as when the donorparent is highly related or markers are used in the selection step.Desired traits that may be used include those nucleic acids known in theart, some of which are listed herein, that will affect traits throughnucleic acid expression or inhibition. Desired loci include theintrogression of FRT, Lox, and other sites for site specificintegration, which may also affect a desired trait if a functionalnucleic acid is inserted at the integration site.

In addition, the above process and other similar processes describedherein may be used to produce first generation progeny soybean seed byadding a step at the end of the process that comprises crossing soybeancultivar 2163154 with the introgressed trait or locus with a differentsoybean plant and harvesting the resultant first-generation progenysoybean seed.

Molecular Techniques Using Soybean Cultivar 2163154

The advent of new molecular biological techniques has allowed theisolation and characterization of genetic elements with specificfunctions, such as encoding specific protein products. Scientists in thefield of plant biology developed a strong interest in engineering thegenome of plants to contain and express foreign genetic elements, oradditional, or modified versions of native or endogenous geneticelements in order to “alter” (the utilization of up-regulation,down-regulation, or gene silencing) the traits of a plant in a specificmanner. Any DNA sequences, whether from a different species or from thesame species, which are introduced into the genome using transformationor various breeding methods are referred to herein collectively as“transgenes.” In some embodiments, a transgenic variant of soybeancultivar 2163154 may contain at least one transgene but could contain atleast 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, and/or no more than 15, 14, 13, 12,11, 10, 9, 8, 7, 6, 5, 4, 3, or 2. Over the last fifteen to twenty yearsseveral methods for producing transgenic plants have been developed, andanother embodiment also relates to transgenic variants of the claimedsoybean variety 2163154.

Nucleic acids or polynucleotides refer to RNA or DNA that is linear orbranched, single or double stranded, or a hybrid thereof. The term alsoencompasses RNA/DNA hybrids. These terms also encompass untranslatedsequence located at both the 3′ and 5′ ends of the coding region of thegene: at least about 1000 nucleotides of sequence upstream from the 5′end of the coding region and at least about 200 nucleotides of sequencedownstream from the 3′ end of the coding region of the gene. Less commonbases, such as inosine, 5-methylcytosine, 6-methyladenine, hypoxanthineand others can also be used for antisense, dsRNA and ribozyme pairing.For example, polynucleotides that contain C-5 propyne analogues ofuridine and cytidine have been shown to bind RNA with high affinity andto be potent antisense inhibitors of gene expression. Othermodifications, such as modification to the phosphodiester backbone, orthe 2′-hydroxy in the ribose sugar group of the RNA can also be made.The antisense polynucleotides and ribozymes can consist entirely ofribonucleotides, or can contain mixed ribonucleotides anddeoxyribonucleotides. The polynucleotides of the embodiments may beproduced by any means, including genomic preparations, cDNApreparations, in-vitro synthesis, RT-PCR, and in vitro or in vivotranscription.

One embodiment is a process for producing soybean variety 2163154further comprising a desired trait, said process comprising introducinga transgene that confers a desired trait to a soybean plant of variety2163154. Another embodiment is the product produced by this process. Inone embodiment, the desired trait may be one or more of herbicideresistance, insect resistance, disease resistance, decreased phytate, ormodified fatty acid or carbohydrate metabolism. The specific gene may beany known in the art or listed herein, including: a polynucleotideconferring resistance to imidazolinone, dicamba, sulfonylurea,glyphosate, glufosinate, triazine, PPO-inhibitor herbicides,benzonitrile, cyclohexanedione, phenoxy proprionic acid, andL-phosphinothricin; a polynucleotide encoding a Bacillus thuringiensispolypeptide; a polynucleotide encoding phytase, FAD-2, FAD-3, galactinolsynthase, or a raffinose synthetic enzyme; or a polynucleotideconferring resistance to soybean cyst nematode, brown stem rot,Phytophthora root rot, soybean mosaic virus, or sudden death syndrome.

Numerous methods for plant transformation have been developed, includingbiological and physical plant transformation protocols. See, forexample, Miki et al., “Procedures for Introducing Foreign DNA intoPlants,” in Methods in Plant Molecular Biology and Biotechnology, Glickand Thompson Eds., CRC Press, Inc., Boca Raton, pp. 67-88 (1993), andArmstrong, “The First Decade of Maize Transformation: A Review andFuture Perspective,” Maydica, 44:101-109 (1999). In addition, expressionvectors and in vitro culture methods for plant cell or tissuetransformation and regeneration of plants are available. See, forexample, Gruber, et al., “Vectors for Plant Transformation,” in Methodsin Plant Molecular Biology and Biotechnology, Glick and Thompson Eds.,CRC Press, Inc., Boca Raton, pp. 89-119 (1993).

A genetic trait which has been engineered into the genome of aparticular soybean plant may then be moved into the genome of anothervariety using traditional breeding techniques that are well known in theplant breeding arts. For example, a backcrossing approach is commonlyused to move a transgene from a transformed soybean variety into analready developed soybean variety, and the resulting backcrossconversion plant would then comprise the transgene(s).

Various genetic elements can be introduced into the plant genome usingtransformation. These elements include, but are not limited to, genes,coding sequences, inducible, constitutive and tissue specific promoters,enhancing sequences, and signal and targeting sequences. For example,see the traits, genes, and transformation methods listed in U.S. Pat.No. 6,118,055.

Breeding with Molecular Markers

Molecular markers, which includes markers identified through the use oftechniques such as Isozyme Electrophoresis, Restriction Fragment LengthPolymorphisms (RFLPs), Randomly Amplified Polymorphic DNAs (RAPDs),Arbitrarily Primed Polymerase Chain Reaction (AP-PCR), DNA AmplificationFingerprinting (DAF), Sequence Characterized Amplified Regions (SCARs),Amplified Fragment Length Polymorphisms (AFLPs), Simple Sequence Repeats(SSRs), and Single Nucleotide Polymorphisms (SNPs), may be used in plantbreeding methods utilizing soybean cultivar 2163154.

Isozyme Electrophoresis and RFLPs have been widely used to determinegenetic composition. Shoemaker and Olsen, Molecular Linkage Map ofSoybean (Glycine max L. Men.), pp. 6.131-6.138 (1993). In S. J. O'Brien(ed.), Genetic Maps: Locus Maps of Complex Genomes, Cold Spring HarborLaboratory Press, Cold Spring Harbor, N.Y., developed a moleculargenetic linkage map that consisted of 25 linkage groups with about 365RFLP, 11 RAPD (random amplified polymorphic DNA), 3 classical markers,and 4 isozyme loci. See also, Shoemaker, R. C., 1994 RFLP Map ofSoybean, pp. 299-309; In R. L. Phillips and I. K. Vasil (ed.), DNA-basedmarkers in plants, Kluwer Academic Press Dordrecht, the Netherlands.

SSR technology is currently the most efficient and practical markertechnology. More marker loci can be routinely used, and more alleles permarker locus can be found, using SSRs in comparison to RFLPs. Forexample, Diwan and Cregan described highly polymorphic microsatelliteloci in soybean with as many as 26 alleles. (Diwan, N., and Cregan. P.B., Automated sizing of fluorescent-labeled simple sequence repeat (SSR)markers to assay genetic variation in Soybean, Theor. Appl. Genet.,95:220-225 (1997)). Single Nucleotide Polymorphisms may also be used toidentify the unique genetic composition of the embodiment(s) and progenyvarieties retaining that unique genetic composition. Various molecularmarker techniques may be used in combination to enhance overallresolution.

Soybean DNA molecular marker linkage maps have been rapidly constructedand widely implemented in genetic studies. One such study is describedin Cregan, et. al, “An Integrated Genetic Linkage Map of the SoybeanGenome,” Crop Science, 39:1464-1490 (1999). Sequences and PCR conditionsof SSR Loci in Soybean, as well as the most current genetic map, may befound in Soybase on the World Wide Web.

One use of molecular markers is Quantitative Trait Loci (QTL) mapping.QTL mapping is the use of markers, which are known to be closely linkedto alleles that have measurable effects on a quantitative trait.Selection in the breeding process is based upon the accumulation ofmarkers linked to the positive effecting alleles and/or the eliminationof the markers linked to the negative effecting alleles from the plant'sgenome.

Molecular markers can also be used during the breeding process for theselection of qualitative traits. For example, markers closely linked toalleles or markers containing sequences within the actual alleles ofinterest can be used to select plants that contain the alleles ofinterest during a backcrossing breeding program. The markers can also beused to select for the genome of the recurrent parent and against thegenome of the donor parent. Using this procedure can minimize the amountof genome from the donor parent that remains in the selected plants. Itcan also be used to reduce the number of crosses back to the recurrentparent needed in a backcrossing program. The use of molecular markers inthe selection process is often called genetic marker enhanced selection.Molecular markers may also be used to identify and exclude certainsources of germplasm as parental varieties or ancestors of a plant byproviding a means of tracking genetic profiles through crosses.

Production of Double Haploids

The production of double haploids can also be used for the developmentof plants with a homozygous phenotype in the breeding program. Forexample, a soybean plant for which soybean cultivar 2163154 is a parentcan be used to produce double haploid plants. Double haploids areproduced by the doubling of a set of chromosomes (1N) from aheterozygous plant to produce a completely homozygous individual. Forexample, see, Wan, et al., “Efficient Production of Doubled HaploidPlants Through Colchicine Treatment of Anther-Derived Maize Callus,”Theoretical and Applied Genetics, 77:889-892 (1989) and U.S. Pat. No.7,135,615. This can be advantageous because the process omits thegenerations of selfing needed to obtain a homozygous plant from aheterozygous source.

Haploid induction systems have been developed for various plants toproduce haploid tissues, plants and seeds. The haploid induction systemcan produce haploid plants from any genotype by crossing a selected line(as female) with an inducer line. Such inducer lines for maize includeStock 6 (Coe, Am. Nat., 93:381-382 (1959); Sharkar and Coe, Genetics,54:453-464 (1966); KEMS (Deimling, Roeber, and Geiger, Vortr.Pflanzenzuchtg, 38:203-224 (1997); or KMS and ZMS (Chalyk, Bylich &Chebotar, MNL, 68:47 (1994); Chalyk & Chebotar, Plant Breeding,119:363-364 (2000)); and indeterminate gametophyte (ig) mutation(Kermicle, Science, 166:1422-1424 (1969)). The disclosures of which areincorporated herein by reference.

Methods for obtaining haploid plants are also disclosed in Kobayashi,M., et al., Journ. of Heredity, 71(1):9-14 (1980); Pollacsek, M.,Agronomie (Paris) 12(3):247-251 (1992); Cho-Un-Haing, et al., Journ. ofPlant Biol., 39(3):185-188 (1996); Verdoodt, L., et al., 96(2):294-300(February 1998); Chalyk, et al., Maize Genet Coop., Newsletter 68:47(1994).

Thus, an embodiment is a process for making a substantially homozygoussoybean cultivar 2163154 progeny plant by producing or obtaining a seedfrom the cross of soybean cultivar 2163154 and another soybean plant andapplying double haploid methods to the F₁ seed or F₁ plant or to anysuccessive filial generation. Based on studies in maize and currentlybeing conducted in soybean, such methods would decrease the number ofgenerations required to produce a variety with similar genetics orcharacteristics to soybean cultivar 2163154. See, Bernardo, R. andKahler, A. L., Theor. Appl. Genet., 102:986-992 (2001).

In particular, a process of making seed retaining the molecular markerprofile of soybean variety 2163154 is contemplated, such processcomprising obtaining or producing F₁ seed for which soybean variety2163154 is a parent, inducing doubled haploids to create progeny withoutthe occurrence of meiotic segregation, obtaining the molecular markerprofile of soybean variety 2163154, and selecting progeny that retainthe molecular marker profile of soybean cultivar 2163154.

Descriptions of other breeding methods that are commonly used fordifferent traits and crops can be found in one of several referencebooks (e.g., Allard (1960); Simmonds (1979); Sneep, et al. (1979); Fehr(1987))

Expression Vectors for Soybean Transformation: Marker Genes

Plant transformation involves the construction of an expression vectorwhich will function in plant cells. Such a vector comprises DNAcomprising a gene under control of, or operatively linked to, aregulatory element (for example, a promoter). Expression vectors includeat least one genetic marker operably linked to a regulatory element (forexample, a promoter) that allows transformed cells containing the markerto be either recovered by negative selection, i.e., inhibiting growth ofcells that do not contain the selectable marker gene, or by positiveselection, i.e., screening for the product encoded by the geneticmarker. Many commonly used selectable marker genes for planttransformation are well-known in the transformation arts, and include,for example, genes that code for enzymes that metabolically detoxify aselective chemical agent which may be an antibiotic or an herbicide, orgenes that encode an altered target which is insensitive to theinhibitor. A few positive selection methods are also known in the art.

One commonly used selectable marker gene for plant transformation is theneomycin phosphotransferase II (nptII) gene which, when under thecontrol of plant regulatory signals, confers resistance to kanamycin.Fraley, et al., Proc. Natl. Acad. Sci. USA, 80:4803 (1983).

Another commonly used selectable marker gene is the hygromycinphosphotransferase gene which confers resistance to the antibiotichygromycin. Vanden Elzen, et al., Plant Mol. Biol., 5:299 (1985).

Additional selectable marker genes of bacterial origin that conferresistance to antibiotics include gentamycin acetyl transferase,streptomycin phosphotransferase and aminoglycoside-3′-adenyltransferase, the bleomycin resistance determinant (Hayford, et al.,Plant Physiol., 86:1216 (1988); Jones, et al., Mol. Gen. Genet., 210:86(1987); Svab, et al., Plant Mol. Biol., 14:197 (1990); Hille, et al.,Plant Mol. Biol., 7:171 (1986)). Other selectable marker genes conferresistance to herbicides such as glyphosate, glufosinate, or bromoxynil(Comai, et al., Nature, 317:741-744 (1985); Gordon-Kamm, et al., PlantCell, 2:603-618 (1990); Stalker, et al., Science, 242:419-423 (1988)).

Selectable marker genes for plant transformation not of bacterial origininclude, for example, mouse dihydrofolate reductase, plant5-enolpyruvylshikimate-3-phosphate synthase, and plant acetolactatesynthase (Eichholtz, et al., Somatic Cell Mol. Genet., 13:67 (1987);Shah, et al., Science, 233:478 (1986); Charest, et al., Plant Cell Rep.,8:643 (1990)).

Another class of marker genes for plant transformation requiresscreening of presumptively transformed plant cells, rather than directgenetic selection of transformed cells, for resistance to a toxicsubstance such as an antibiotic. These genes are particularly useful toquantify or visualize the spatial pattern of expression of a gene inspecific tissues and are frequently referred to as reporter genesbecause they can be fused to a gene or gene regulatory sequence for theinvestigation of gene expression. Commonly used genes for screeningpresumptively transformed cells include β-glucuronidase (GUS),β-galactosidase, luciferase, and chloramphenicol acetyltransferase(Jefferson, R. A., Plant Mol. Biol. Rep., 5:387 (1987); Teeri, et al.,EMBO J., 8:343 (1989); Koncz, et al., Proc. Natl. Acad. Sci. USA, 84:131(1987); DeBlock, et al., EMBO J., 3:1681 (1984)).

In vivo methods for visualizing GUS activity that do not requiredestruction of plant tissue are available (Molecular Probes, Publication2908, IMAGENE GREEN, pp. 1-4 (1993); Naleway, et al., J. Cell Biol.,115:151a (1991)). However, these in-vivo methods for visualizing GUSactivity have not proven useful for recovery of transformed cellsbecause of low sensitivity, high fluorescent backgrounds, andlimitations associated with the use of luciferase genes as selectablemarkers.

More recently, a gene encoding Green Fluorescent Protein (GFP) has beenutilized as a marker for gene expression in prokaryotic and eukaryoticcells (Chalfie, et al., Science, 263:802 (1994)). GFP and mutants of GFPmay be used as screenable markers.

Expression Vectors for Soybean Transformation: Promoters

Genes included in expression vectors must be driven by a nucleotidesequence comprising a regulatory element (for example, a promoter).Several types of promoters are well known in the transformation arts asare other regulatory elements that can be used alone or in combinationwith promoters.

As used herein, “promoter” includes reference to a region of DNAupstream from the start of transcription and involved in recognition andbinding of RNA polymerase and other proteins to initiate transcription.A “plant promoter” is a promoter capable of initiating transcription inplant cells. Examples of promoters under developmental control includepromoters that preferentially initiate transcription in certain tissues,such as leaves, roots, seeds, fibers, xylem vessels, tracheids, orsclerenchyma. Such promoters are referred to as “tissue-preferred.”Promoters that initiate transcription only in a certain tissue arereferred to as “tissue-specific.” A “cell-type” specific promoterprimarily drives expression in certain cell types in one or more organs,for example, vascular cells in roots or leaves. An “inducible” promoteris a promoter which is under environmental control. Examples ofenvironmental conditions that may affect transcription by induciblepromoters include anaerobic conditions or the presence of light.Tissue-specific, tissue-preferred, cell-type specific, and induciblepromoters constitute the class of “non-constitutive” promoters. A“constitutive” promoter is a promoter that is active under mostenvironmental conditions.

A. Inducible Promoters: An inducible promoter is operably linked to agene for expression in soybean. Optionally, the inducible promoter isoperably linked to a nucleotide sequence encoding a signal sequencewhich is operably linked to a gene for expression in soybean. With aninducible promoter, the rate of transcription increases in response toan inducing agent.

Any inducible promoter can be used in an embodiment(s). See, Ward, etal., Plant Mol. Biol., 22:361-366 (1993). Exemplary inducible promotersinclude, but are not limited to, that from the ACEI system whichresponds to copper (Mett, et al., Proc. Natl. Acad. Sci. USA,90:4567-4571 (1993)); In2 gene from maize which responds tobenzenesulfonamide herbicide safeners (Hershey, et al., Mol. GenGenetics, 227:229-237 (1991); Gatz, et al., Mol. Gen. Genetics,243:32-38 (1994)); or Tet repressor from Tn10 (Gatz, et al., Mol. Gen.Genetics, 227:229-237 (1991)). An inducible promoter is a promoter thatresponds to an inducing agent to which plants do not normally respond.An exemplary inducible promoter is the inducible promoter from a steroidhormone gene, the transcriptional activity of which is induced by aglucocorticosteroid hormone (Schena, et al., Proc. Natl. Acad. Sci. USA,88:0421 (1991)).

B. Constitutive Promoters: A constitutive promoter is operably linked toa gene for expression in soybean or the constitutive promoter isoperably linked to a nucleotide sequence encoding a signal sequencewhich is operably linked to a gene for expression in soybean.

Many different constitutive promoters can be utilized in anembodiment(s). Exemplary constitutive promoters include, but are notlimited to, the promoters from plant viruses such as the 35S promoterfrom CaMV (Odell, et al., Nature, 313:810-812 (1985)) and the promotersfrom such genes as rice actin (McElroy, et al., Plant Cell, 2:163-171(1990)); ubiquitin (Christensen, et al., Plant Mol. Biol., 12:619-632(1989); Christensen, et al., Plant Mol. Biol., 18:675-689 (1992)); pEMU(Last, et al., Theor. Appl. Genet., 81:581-588 (1991)); MAS (Velten, etal., EMBO J., 3:2723-2730 (1984)); and maize H3 histone (Lepetit, etal., Mol. Gen. Genetics, 231:276-285 (1992); Atanassova, et al., PlantJournal, 2 (3):291-300 (1992)). The ALS promoter, Xbal/NcoI fragment 5′to the Brassica napus ALS3 structural gene (or a nucleotide sequencesimilarity to said XbaI/NcoI fragment), represents a particularly usefulconstitutive promoter. See, U.S. Pat. No. 5,659,026.

C. Tissue-Specific or Tissue-Preferred Promoters: A tissue-specificpromoter is operably linked to a gene for expression in soybean.Optionally, the tissue-specific promoter is operably linked to anucleotide sequence encoding a signal sequence which is operably linkedto a gene for expression in soybean. Plants transformed with a gene ofinterest operably linked to a tissue-specific promoter produce theprotein product of the transgene exclusively, or preferentially, in aspecific tissue.

Any tissue-specific or tissue-preferred promoter can be utilized in anembodiment(s). Exemplary tissue-specific or tissue-preferred promotersinclude, but are not limited to, a root-preferred promoter such as thatfrom the phaseolin gene (Murai, et al., Science, 23:476-482 (1983);Sengupta-Gopalan, et al., Proc. Natl. Acad. Sci. USA, 82:3320-3324(1985)); a leaf-specific and light-induced promoter such as that fromcab or rubisco (Simpson, et al., EMBO J., 4(11):2723-2729 (1985); Timko,et al., Nature, 318:579-582 (1985)); an anther-specific promoter such asthat from LAT52 (Twell, et al., Mol. Gen. Genetics, 217:240-245 (1989));a pollen-specific promoter such as that from Zm13 (Guerrero, et al.,Mol. Gen. Genetics, 244:161-168 (1993)); or a microspore-preferredpromoter such as that from apg (Twell, et al., Sex. Plant Reprod.,6:217-224 (1993)).

Signal Sequences for Targeting Proteins to Subcellular Compartments

Transport of a protein produced by transgenes to a subcellularcompartment, such as the chloroplast, vacuole, peroxisome, glyoxysome,cell wall, or mitochondrion, or for secretion into the apoplast, isaccomplished by means of operably linking the nucleotide sequenceencoding a signal sequence to the 5′ and/or 3′ region of a gene encodingthe protein of interest. Targeting sequences at the 5′ and/or 3′ end ofthe structural gene may determine during protein synthesis andprocessing where the encoded protein is ultimately compartmentalized.

The presence of a signal sequence directs a polypeptide to either anintracellular organelle or subcellular compartment or for secretion tothe apoplast. Many signal sequences are known in the art. See, forexample, Becker, et al., Plant Mol. Biol., 20:49 (1992); Knox, C., etal., Plant Mol. Biol., 9:3-17 (1987); Lerner, et al., Plant Physiol.,91:124-129 (1989); Frontes, et al., Plant Cell, 3:483-496 (1991);Matsuoka, et al., Proc. Natl. Acad. Sci., 88:834 (1991); Gould, et al.,J. Cell. Biol., 108:1657 (1989); Creissen, et al., Plant J., 2:129(1991); Kalderon, et al., Cell, 39:499-509 (1984); Steifel, et al.,Plant Cell, 2:785-793 (1990).

Foreign Protein Genes and Agronomic Genes: Transformation

With transgenic plants according to one embodiment, a foreign proteincan be produced in commercial quantities. Thus, techniques for theselection and propagation of transformed plants, which are wellunderstood in the art, yield a plurality of transgenic plants which areharvested in a conventional manner, and a foreign protein can then beextracted from a tissue of interest or from total biomass. Proteinextraction from plant biomass can be accomplished by known methods whichare discussed, for example, by Heney and Orr, Anal. Biochem., 114:92-6(1981).

According to an embodiment, the transgenic plant provided for commercialproduction of foreign protein is a soybean plant. In another embodiment,the biomass of interest is seed. For the relatively small number oftransgenic plants that show higher levels of expression, a genetic mapcan be generated, primarily via conventional RFLP, PCR, and SSRanalysis, which identifies the approximate chromosomal location of theintegrated DNA molecule. For exemplary methodologies in this regard,see, Glick and Thompson, Methods in Plant Molecular Biology andBiotechnology, CRC Press, Inc., Boca Raton, 269:284 (1993). Mapinformation concerning chromosomal location is useful for proprietaryprotection of a subject transgenic plant.

Wang, et al. discuss “Large Scale Identification, Mapping and Genotypingof Single-Nucleotide Polymorphisms in the Human Genome,” Science,280:1077-1082 (1998), and similar capabilities are becoming increasinglyavailable for the soybean genome. Map information concerning chromosomallocation is useful for proprietary protection of a subject transgenicplant. If unauthorized propagation is undertaken and crosses made withother germplasm, the map of the integration region can be compared tosimilar maps for suspect plants to determine if the latter have a commonparentage with the subject plant. Map comparisons would involvehybridizations, RFLP, PCR, SSR, and sequencing, all of which areconventional techniques. SNPs may also be used alone or in combinationwith other techniques.

Likewise, by means of one embodiment, plants can be geneticallyengineered to express various phenotypes of agronomic interest. Throughthe transformation of soybean, the expression of genes can be altered toenhance disease resistance, insect resistance, herbicide resistance,agronomic, grain quality, and other traits. Transformation can also beused to insert DNA sequences which control or help controlmale-sterility. DNA sequences native to soybean, as well as non-nativeDNA sequences, can be transformed into soybean and used to alter levelsof native or non-native proteins. Various promoters, targetingsequences, enhancing sequences, and other DNA sequences can be insertedinto the genome for the purpose of altering the expression of proteins.The interruption or suppression of the expression of a gene at the levelof transcription or translation (also known as gene silencing or genesuppression) is desirable for several aspects of genetic engineering inplants.

Many techniques for gene silencing are well-known to one of skill in theart, including, but not limited to, knock-outs (such as by insertion ofa transposable element such as Mu (Vicki Chandler, The Maize Handbook,Ch. 118 (Springer-Verlag 1994)) or other genetic elements such as a FRT,Lox, or other site specific integration sites; antisense technology(see, e.g., Sheehy, et al., PNAS USA, 85:8805-8809 (1988) and U.S. Pat.Nos. 5,107,065, 5,453,566, and 5,759,829); co-suppression (e.g., Taylor,Plant Cell, 9:1245 (1997); Jorgensen, Trends Biotech., 8(12):340-344(1990); Flavell, PNAS USA, 91:3490-3496 (1994); Finnegan, et al.,Bio/Technology, 12:883-888 (1994); Neuhuber, et al., Mol. Gen. Genet.,244:230-241 (1994)); RNA interference (Napoli, et al., Plant Cell,2:279-289 (1990); U.S. Pat. No. 5,034,323; Sharp, Genes Dev., 13:139-141(1999); Zamore, et al., Cell, 101:25-33 (2000); Montgomery, et al., PNASUSA, 95:15502-15507 (1998)), virus-induced gene silencing (Burton, etal., Plant Cell, 12:691-705 (2000); Baulcombe, Curr. Op. Plant Bio.,2:109-113 (1999)); target-RNA-specific ribozymes (Haseloff, et al.,Nature, 334:585-591 (1988)); hairpin structures (Smith, et al., Nature,407:319-320 (2000); U.S. Pat. Nos. 6,423,885, 7,138,565, 6,753,139, and7,713,715); MicroRNA (Aukerman & Sakai, Plant Cell, 15:2730-2741(2003)); ribozymes (Steinecke, et al., EMBO J., 11:1525 (1992);Perriman, et al., Antisense Res. Dev., 3:253 (1993)); oligonucleotidemediated targeted modification (e.g., U.S. Pat. Nos. 6,528,700 and6,911,575); Zn-finger targeted molecules (e.g., U.S. Pat. Nos.7,151,201, 6,453,242, 6,785,613, 7,177,766 and 7,788,044); and othermethods or combinations of the above methods known to those of skill inthe art.

Methods for Soybean Transformation

Numerous methods for plant transformation have been developed includingbiological and physical plant transformation protocols. See, forexample, Miki, et al., “Procedures for Introducing Foreign DNA intoPlants,” in Methods in Plant Molecular Biology and Biotechnology, Glickand Thompson Eds., CRC Press, Inc., Boca Raton, pp. 67-88 (1993). Inaddition, expression vectors and in-vitro culture methods for plant cellor tissue transformation and regeneration of plants are available. See,for example, Gruber, et al., “Vectors for Plant Transformation,” inMethods in Plant Molecular Biology and Biotechnology, Glick and ThompsonEds., CRC Press, Inc., Boca Raton, pp. 89-119 (1993).

A. Agrobacterium-mediated Transformation: One method for introducing anexpression vector into plants is based on the natural transformationsystem of Agrobacterium. See, for example, Horsch, et al., Science,227:1229 (1985). A. tumefaciens and A. rhizogenes are plant pathogenicsoil bacteria which genetically transform plant cells. The Ti and Riplasmids of A. tumefaciens and A. rhizogenes, respectively, carry genesresponsible for genetic transformation of the plant. See, for example,Kado, C. I., Crit. Rev. Plant Sci., 10:1 (1991). Descriptions ofAgrobacterium vector systems and methods for Agrobacterium-mediated genetransfer are provided by Gruber, et al., supra, Miki, et al., supra, andMoloney, et al., Plant Cell Reports, 8:238 (1989). See also, U.S. Pat.No. 5,563,055 (Townsend and Thomas), issued Oct. 8, 1996.

B. Direct Gene Transfer: Several methods of plant transformation,collectively referred to as direct gene transfer, have been developed asan alternative to Agrobacterium-mediated transformation. A generallyapplicable method of plant transformation is microprojectile-mediatedtransformation where DNA is carried on the surface of microprojectilesmeasuring 1 to 4 μm. The expression vector is introduced into planttissues with a biolistic device that accelerates the microprojectiles tospeeds of 300 to 600 m/s which is sufficient to penetrate plant cellwalls and membranes. Sanford, et al., Part. Sci. Technol., 5:27 (1987);Sanford, J. C., Trends Biotech., 6:299 (1988); Klein, et al., Bio/Tech.,6:559-563 (1988); Sanford, J. C., Physiol Plant, 7:206 (1990); Klein, etal., Biotechnology, 10:268 (1992). See also, U.S. Pat. No. 5,015,580(Christou, et al.), issued May 14, 1991 and U.S. Pat. No. 5,322,783.

Another method for physical delivery of DNA to plants is sonication oftarget cells. Zhang, et al., Bio/Technology, 9:996 (1991).Alternatively, liposome and spheroplast fusion have been used tointroduce expression vectors into plants. Deshayes, et al., EMBO J.,4:2731 (1985); Christou, et al., Proc Natl. Acad. Sci. USA, 84:3962(1987). Direct uptake of DNA into protoplasts using CaCl₂ precipitation,polyvinyl alcohol or poly-L-ornithine have also been reported. Hain, etal., Mol. Gen. Genet., 199:161 (1985) and Draper, et al., Plant CellPhysiol., 23:451 (1982). Electroporation of protoplasts and whole cellsand tissues has also been described (D'Halluin, et al., Plant Cell,4:1495-1505 (1992); and Spencer, et al., Plant Mol. Biol., 24:51-61(1994)).

Following transformation of soybean target tissues, expression of theabove-described selectable marker genes allows for preferentialselection of transformed cells, tissues, and/or plants, usingregeneration and selection methods well known in the art.

The foregoing methods for transformation would typically be used forproducing a transgenic variety. The transgenic variety could then becrossed with another (non-transformed or transformed) variety in orderto produce a new transgenic variety. Alternatively, a genetic trait thathas been engineered into a particular soybean line using the foregoingtransformation techniques could be moved into another line usingtraditional backcrossing techniques that are well known in the plantbreeding arts. For example, a backcrossing approach could be used tomove an engineered trait from a public, non-elite variety into an elitevariety, or from a variety containing a foreign gene in its genome intoa variety or varieties that do not contain that gene. As used herein,“crossing” can refer to a simple x by y cross or the process ofbackcrossing depending on the context.

Likewise, by means of one embodiment, agronomic genes can be expressedin transformed plants. More particularly, plants can be geneticallyengineered to express various phenotypes of agronomic interest.Exemplary genes implicated in this regard include, but are not limitedto, those categorized below:

1. Genes That Confer Resistance to Pests or Disease and That Encode:

A. Plant disease resistance genes. Plant defenses are often activated byspecific interaction between the product of a disease resistance gene(R) in the plant and the product of a corresponding avirulence (Avr)gene in the pathogen. A plant variety can be transformed with one ormore cloned resistance genes to engineer plants that are resistant tospecific pathogen strains. See, for example, Jones, et al., Science,266:789 (1994) (cloning of the tomato Cf-9 gene for resistance toCladosporium fulvum); Martin, et al., Science, 262:1432 (1993) (tomatoPto gene for resistance to Pseudomonas syringae pv. tomato encodes aprotein kinase); Mindrinos, et al., Cell, 78:1089 (1994) (ArabidopsisRSP2 gene for resistance to Pseudomonas syringae); McDowell & Woffenden,Trends Biotechnol., 21(4):178-83 (2003); and Toyoda, et al., TransgenicRes., 11 (6):567-82 (2002).

B. A gene conferring resistance to a pest, such as soybean cystnematode. See, e.g., U.S. Pat. No. 5,994,627.

C. A Bacillus thuringiensis protein, a derivative thereof or a syntheticpolypeptide modeled thereon. See, for example, Geiser, et al., Gene,48:109 (1986), who disclose the cloning and nucleotide sequence of a Btδ-endotoxin gene. Moreover, DNA molecules encoding δ-endotoxin genes canbe purchased from American Type Culture Collection, Manassas, Va., forexample, under ATCC Accession Nos. 40098, 67136, 31995, and 31998.

D. A lectin. See, for example, Van Damme, et al., Plant Molec. Biol.,24:25 (1994), who disclose the nucleotide sequences of several Cliviaminiata mannose-binding lectin genes.

E. A vitamin-binding protein such as avidin. See, InternationalApplication No. PCT/US1993/006487, which teaches the use of avidin andavidin homologues as larvicides against insect pests.

F. An enzyme inhibitor, for example, a protease or proteinase inhibitoror an amylase inhibitor. See, for example, Abe, et al., J. Biol. Chem.,262:16793 (1987) (nucleotide sequence of rice cysteine proteinaseinhibitor); Huub, et al., Plant Molec. Biol., 21:985 (1993) (nucleotidesequence of cDNA encoding tobacco proteinase inhibitor I); Sumitani, etal., Biosci. Biotech. Biochem., 57:1243 (1993) (nucleotide sequence ofStreptomyces nitrosporeus α-amylase inhibitor); and U.S. Pat. No.5,494,813.

G. An insect-specific hormone or pheromone, such as an ecdysteroid orjuvenile hormone, a variant thereof, a mimetic based thereon, or anantagonist or agonist thereof. See, for example, the disclosure byHammock, et al., Nature, 344:458 (1990), of baculovirus expression ofcloned juvenile hormone esterase, an inactivator of juvenile hormone.

H. An insect-specific peptide or neuropeptide which, upon expression,disrupts the physiology of the affected pest. For example, see thedisclosures of Regan, J. Biol. Chem., 269:9 (1994) (expression cloningyields DNA coding for insect diuretic hormone receptor); Pratt, et al.,Biochem. Biophys. Res. Comm., 163:1243 (1989) (an allostatin isidentified in Diploptera puntata); Chattopadhyay, et al., CriticalReviews in Microbiology, 30(1):33-54 (2004); Zjawiony, J. Nat. Prod.,67(2):300-310 (2004); Carlini & Grossi-de-Sa, Toxicon, 40(11):1515-1539(2002); Ussuf, et al., Curr Sci., 80(7):847-853 (2001); Vasconcelos &Oliveira, Toxicon, 44(4):385-403 (2004). See also, U.S. Pat. No.5,266,317 which discloses genes encoding insect-specific, paralyticneurotoxins.

I. An insect-specific venom produced in nature by a snake, a wasp, etc.For example, see, Pang, et al., Gene, 116:165 (1992), for disclosure ofheterologous expression in plants of a gene coding for a scorpioninsectotoxic peptide.

J. An enzyme responsible for a hyperaccumulation of a monoterpene, asesquiterpene, a steroid, hydroxamic acid, a phenylpropanoid derivative,or another non-protein molecule with insecticidal activity.

K. An enzyme involved in the modification, including thepost-translational modification, of a biologically active molecule; forexample, a glycolytic enzyme, a proteolytic enzyme, a lipolytic enzyme,a nuclease, a cyclase, a transaminase, an esterase, a hydrolase, aphosphatase, a kinase, a phosphorylase, a polymerase, an elastase, achitinase, and a glucanase, whether natural or synthetic. See, U.S. Pat.No. 5,955,653 which discloses the nucleotide sequence of a callase gene.DNA molecules which contain chitinase-encoding sequences can beobtained, for example, from the ATCC under Accession Nos. 39637 and67152. See also, Kramer, et al., Insect Biochem. Molec. Biol., 23:691(1993), who teach the nucleotide sequence of a cDNA encoding tobaccohornworm chitinase, and Kawalleck, et al., Plant Molec. Biol., 21:673(1993), who provide the nucleotide sequence of the parsley ubi4-2polyubiquitin gene, U.S. Pat. Nos. 7,145,060, 7,087,810, and 6,563,020.

L. A molecule that stimulates signal transduction. For example, see thedisclosure by Botella, et al., Plant Molec. Biol., 24:757 (1994), ofnucleotide sequences for mung bean calmodulin cDNA clones, and Griess,et al., Plant Physiol., 104:1467 (1994), who provide the nucleotidesequence of a maize calmodulin cDNA clone.

M. A hydrophobic moment peptide. See, U.S. Pat. No. 5,580,852, whichdiscloses peptide derivatives of tachyplesin which inhibit fungal plantpathogens, and U.S. Pat. No. 5,607,914 which teaches syntheticantimicrobial peptides that confer disease resistance.

N. A membrane permease, a channel former or a channel blocker. Forexample, see the disclosure of Jaynes, et al., Plant Sci, 89:43 (1993),of heterologous expression of a cecropin-13 lytic peptide analog torender transgenic tobacco plants resistant to Pseudomonas solanacearum.

O. A viral-invasive protein or a complex toxin derived therefrom. Forexample, the accumulation of viral coat proteins in transformed plantcells imparts resistance to viral infection and/or disease developmenteffected by the virus from which the coat protein gene is derived, aswell as by related viruses. See, Beachy, et al., Ann. Rev. Phytopathol.,28:451 (1990). Coat protein-mediated resistance has been conferred upontransformed plants against alfalfa mosaic virus, cucumber mosaic virus,and tobacco mosaic virus.

P. An insect-specific antibody or an immunotoxin derived therefrom.Thus, an antibody targeted to a critical metabolic function in theinsect gut would inactivate an affected enzyme, killing the insect.

Q. A virus-specific antibody. See, for example, Tavladoraki, et al.,Nature, 366:469 (1993), who show that transgenic plants expressingrecombinant antibody genes are protected from virus attack.

R. A developmental-arrestive protein produced in nature by a pathogen ora parasite. Thus, fungal endo-α-1,4-D-polygalacturonases facilitatefungal colonization and plant nutrient release by solubilizing plantcell wall homo-α-1,4-D-galacturonase. See, Lamb, et al., Bio/Technology,10:1436 (1992). The cloning and characterization of a gene which encodesa bean endopolygalacturonase-inhibiting protein is described by Toubart,et al., Plant J., 2:367 (1992).

S. A developmental-arrestive protein produced in nature by a plant. Forexample, Logemann, et al., Bio/Technology, 10:305 (1992), have shownthat transgenic plants expressing the barley ribosome-inactivating genehave an increased resistance to fungal disease.

T. Genes involved in the Systemic Acquired Resistance (SAR) Responseand/or the pathogenesis-related genes. Briggs, S., Current Biology, 5(2)(1995); Pieterse & Van Loon, Curr. Opin. Plant Bio., 7(4):456-64 (2004);and Somssich, Cell, 113(7):815-6 (2003).

U. Antifungal genes. See, Cornelissen and Melchers, Plant Physiol.,101:709-712 (1993); Parijs, et al., Planta, 183:258-264 (1991); andBushnell, et al., Can. J of Plant Path., 20(2):137-149 (1998). See also,U.S. Pat. No. 6,875,907.

V. Detoxification genes, such as for fumonisin, beauvericin,moniliformin, and zearalenone and their structurally-relatedderivatives. See, U.S. Pat. No. 5,792,931.

W. Cystatin and cysteine proteinase inhibitors. See, U.S. Pat. No.7,205,453.

X. Defensin genes. See, U.S. Pat. Nos. 6,911,577, 7,855,327, 7,855,328,7,897,847, 7,910,806, 7,919,686, and 8,026,415.

Y. Genes conferring resistance to nematodes, and in particular soybeancyst nematodes. See, U.S. Pat. Nos. 5,994,627 and 6,294,712; Urwin, etal., Planta, 204:472-479 (1998); Williamson, Curr Opin Plant Bio.,2(4):327-31 (1999).

Z. Genes that confer resistance to Phytophthora Root Rot, such as theRps 1, Rps 1-a, Rps 1-b, Rps 1-c, Rps 1-d, Rps 1-e, Rps 1-k, Rps 2, Rps3-a, Rps 3-b, Rps 3-c, Rps 4, Rps 5, Rps 6, Rps 7, and other Rps genes.See, for example, Shoemaker, et al., Phytophthora Root Rot ResistanceGene Mapping in Soybean, Plant Genome IV Conference, San Diego, Calif.(1995).

AA. Genes that confer resistance to Brown Stem Rot, such as described inU.S. Pat. No. 5,689,035 and incorporated by reference for this purpose.

Any of the above-listed disease or pest resistance genes (A-AA) can beintroduced into the claimed soybean cultivar through a variety of meansincluding, but not limited to, transformation and crossing.

2. Genes That Confer Resistance to an Herbicide, for Example:

A. An herbicide that inhibits the growing point or meristem, such as animidazolinone or a sulfonylurea. Exemplary genes in this category codefor mutant ALS and AHAS enzyme as described, for example, by Lee, etal., EMBO J., 7:1241 (1988) and Mild, et al., Theor. Appl. Genet.,80:449 (1990), respectively.

B. Glyphosate (resistance conferred by mutant5-enolpyruvlshikimate-3-phosphate synthase (EPSPS) and aroA genes,respectively) and other phosphono compounds, such as glufosinate(phosphinothricin acetyl transferase (PAT) and Streptomyceshygroscopicus PAT bar genes), pyridinoxy or phenoxy proprionic acids,and cyclohexanediones (ACCase inhibitor-encoding genes). See, forexample, U.S. Pat. No. 4,940,835 which discloses the nucleotide sequenceof a form of EPSPS which can confer glyphosate resistance. U.S. Pat. No.5,627,061 which describes genes encoding EPSPS enzymes. See also, U.S.Pat. Nos. 6,566,587, 6,338,961, 6,248,876, 6,040,497, 5,804,425,5,633,435, 5,145,783, 4,971,908, 5,312,910, 5,188,642, 4,940,835,5,866,775, 6,225,114, 6,130,366, 5,310,667, 4,535,060, 4,769,061,5,633,448, 5,510,471, 6,803,501, RE 36,449, RE 37,287, and U.S. Pat. No.5,491,288, which are incorporated herein by reference for this purpose.Glyphosate resistance is also imparted to plants that express a genethat encodes a glyphosate oxido-reductase enzyme, as described morefully in U.S. Pat. Nos. 5,776,760 and 5,463,175, which are incorporatedherein by reference for this purpose. In addition, glyphosate resistancecan be imparted to plants by the over expression of genes encodingglyphosate N-acetyltransferase. See, for example, U.S. Pat. No.7,462,481. A DNA molecule encoding a mutant aroA gene can be obtainedunder ATCC Accession No. 39256, and the nucleotide sequence of themutant gene is disclosed in U.S. Pat. No. 4,769,061. European PatentAppl. No. 0333033 and U.S. Pat. No. 4,975,374 disclose nucleotidesequences of glutamine synthetase genes which confer resistance toherbicides such as L-phosphinothricin. The nucleotide sequence of a PATgene is provided in European Patent No. 0242246 to Leemans, et al.DeGreef, et al., Bio/Technology, 7:61 (1989) describe the production oftransgenic plants that express chimeric bar genes coding forphosphinothricin acetyl transferase activity. Exemplary of genesconferring resistance to phenoxy proprionic acids and cyclohexones, suchas sethoxydim and haloxyfop are the Acc1-S1, Acc1-S2, and Acc2-S3 genesdescribed by Marshall, et al., Theor. Appl. Genet., 83:435 (1992).

C. An herbicide that inhibits photosynthesis, such as a triazine (psbAand gs+ genes) and a benzonitrile (nitrilase gene). Przibila, et al.,Plant Cell, 3:169 (1991), describe the transformation of Chlamydomonaswith plasmids encoding mutant psbA genes. Nucleotide sequences fornitrilase genes are disclosed in U.S. Pat. No. 4,810,648 to Stalker andDNA molecules containing these genes are available under ATCC AccessionNos. 53435, 67441, and 67442. Cloning and expression of DNA coding for aglutathione S-transferase is described by Hayes, et al., Biochem. J.,285:173 (1992). Protoporphyrinogen oxidase (PPO) is the target of thePPO-inhibitor class of herbicides; a PPO-inhibitor resistant PPO genewas recently identified in Amaranthus tuberculatus (Patzoldt et al.,PNAS, 103(33):12329-2334, 2006). The herbicide methyl viologen inhibitsCO₂ assimilation. Foyer et al. (Plant Physiol., 109:1047-1057, 1995)describe a plant overexpressing glutathione reductase (GR) which isresistant to methyl viologen treatment. Bromoxynil resistance byintroducing a chimeric gene containing the bxn gene (Science, 242(4877):419-23, 1988).

D. Acetohydroxy acid synthase, which has been found to make plants thatexpress this enzyme resistant to multiple types of herbicides, has beenintroduced into a variety of plants. See, Hattori, et al., Mol. Gen.Genet., 246:419 (1995). Other genes that confer tolerance to herbicidesinclude a gene encoding a chimeric protein of rat cytochrome P4507A1 andyeast NADPH-cytochrome P450 oxidoreductase (Shiota, et al., PlantPhysiol., 106:17 (1994)); genes for glutathione reductase and superoxidedismutase (Aono, et al., Plant Cell Physiol., 36:1687 (1995)); and genesfor various phosphotransferases (Datta, et al., Plant Mol. Biol., 20:619(1992)).

E. Protoporphyrinogen oxidase (protox) is necessary for the productionof chlorophyll, which is necessary for all plant survival. The protoxenzyme serves as the target for a variety of herbicidal compounds. Theseherbicides also inhibit growth of all the different species of plantspresent, causing their total destruction. The development of plantscontaining altered protox activity which are resistant to theseherbicides are described in U.S. Pat. Nos. 6,288,306, 6,282,837,5,767,373, and 6,084,155.

Any of the above listed herbicide genes (A-E) can be introduced into theclaimed soybean cultivar through a variety of means including but notlimited to transformation and crossing.

3. Genes That Confer or Contribute to a Value-Added Trait, such as:

A. Modified fatty acid metabolism, for example, by transforming a plantwith an antisense gene of stearyl-ACP desaturase to increase stearicacid content of the plant. See, Knultzon, et al., Proc. Natl. Acad. Sci.USA, 89:2625 (1992).

B. Decreased phytate content: 1) Introduction of a phytase-encoding geneenhances breakdown of phytate, adding more free phosphate to thetransformed plant. For example, see, Van Hartingsveldt, et al., Gene,127:87 (1993), for a disclosure of the nucleotide sequence of anAspergillus niger phytase gene. 2) Up-regulation of a gene that reducesphytate content. In maize, this, for example, could be accomplished bycloning and then re-introducing DNA associated with one or more of thealleles, such as the LPA alleles, identified in maize mutantscharacterized by low levels of phytic acid, such as in Raboy, et al.,Maydica, 35:383 (1990), and/or by altering inositol kinase activity asin, for example, U.S. Pat. Nos. 7,425,442, 7,714,187, 6,197,561,6,2191,224, 6,855,869, 6,391,348, 6,197,561, and 6,291,224; U.S. Publ.Nos. 2003/000901, 2003/0009011, and 2006/272046; and International Pub.Nos. WO 98/45448, and WO 01/04147.

C. Modified carbohydrate composition effected, for example, bytransforming plants with a gene coding for an enzyme that alters thebranching pattern of starch, or a gene altering thioredoxin, such as NTRand/or TRX (See, U.S. Pat. No. 6,531,648, which is incorporated byreference for this purpose), and/or a gamma zein knock out or mutant,such as cs27 or TUSC27 or en27 (See, U.S. Pat. Nos. 6,858,778, 7,741,533and U.S. Publ. No. 2005/0160488, which are incorporated by reference forthis purpose). See, Shiroza, et al., J. Bacteriol., 170:810 (1988)(nucleotide sequence of Streptococcus mutans fructosyltransferase gene);Steinmetz, et al., Mol. Gen. Genet., 200:220 (1985) (nucleotide sequenceof Bacillus subtilis levansucrase gene); Pen, et al., Bio/Technology,10:292 (1992) (production of transgenic plants that express Bacilluslicheniformis α-amylase); Elliot, et al., Plant Molec. Biol., 21:515(1993) (nucleotide sequences of tomato invertase genes); Søgaard, etal., J. Biol. Chem., 268:22480-22484 (1993) (site-directed mutagenesisof barley α-amylase gene); Fisher, et al., Plant Physiol., 102:1045(1993) (maize endosperm starch branching enzyme II); International Pub.No. WO 99/10498 (improved digestibility and/or starch extraction throughmodification of UDP-D-xylose 4-epimerase, Fragile 1 and 2, Ref 1, HCHL,C4H); U.S. Pat. No. 6,232,529 (method of producing high oil seed bymodification of starch levels (AGP)). The fatty acid modification genesmentioned above may also be used to affect starch content and/orcomposition through the interrelationship of the starch and oilpathways.

D. Elevated oleic acid via FAD-2 gene modification and/or decreasedlinolenic acid via FAD-3 gene modification. See, U.S. Pat. Nos.5,952,544, 6,063,947, and 6,323,392. Linolenic acid is one of the fivemost abundant fatty acids in soybean seeds. The low oxidative stabilityof linolenic acid is one reason that soybean oil undergoes partialhydrogenation. When partially hydrogenated, all unsaturated fatty acidsform trans fats. Soybeans are the largest source of edible-oils in theU.S. and 40% of soybean oil production is partially hydrogenated. Theconsumption of trans fats increases the risk of heart disease.Regulations banning trans fats have encouraged the development of lowlinolenic soybeans. Soybeans containing low linolenic acid percentagescreate a more stable oil requiring hydrogenation less often. Thisprovides trans fat free alternatives in products such as cooking oil.

E. Altering conjugated linolenic or linoleic acid content, such as inU.S. Pat. No. 6,593,514. Altering LEC1, AGP, Dek1, Superal1, milps, andvarious Ipa genes, such as Ipa1, Ipa3, hpt, or hggt. See, for example,U.S. Pat. Nos. 7,122,658, 7,342,418, 6,232,529, 7,888,560, 6,423,886,6,197,561, 6,825,397 and 7,157,621; U.S. Publ. No. 2003/0079247;International Publ. No. WO 2003/011015; and Rivera-Madrid, R., et al.,Proc. Natl. Acad. Sci., 92:5620-5624 (1995).

F. Altered antioxidant content or composition, such as alteration oftocopherol or tocotrienols. See, for example, U.S. Pat. Nos. 6,787,683,7,154,029 and International Publ. No. WO 00/68393 (involving themanipulation of antioxidant levels through alteration of a phyt1 prenyltransferase (ppt)); and U.S. Pat. Nos. 7,154,029 and 7,622,658 (throughalteration of a homogentisate geranyl geranyl transferase (hggt)).

G. Altered essential seed amino acids. See, for example, U.S. Pat. No.6,127,600 (method of increasing accumulation of essential amino acids inseeds); U.S. Pat. No. 6,080,913 (binary methods of increasingaccumulation of essential amino acids in seeds); U.S. Pat. No. 5,990,389and International Publ. No. WO 95/15392 (high lysine); U.S. Pat. No.5,850,016 (alteration of amino acid compositions in seeds); U.S. Pat.No. 5,885,802 (high methionine); U.S. Pat. No. 5,885,801 andInternational Publ. No. WO96/01905 (high threonine); U.S. Pat. No.6,664,445, 7,022,895, 7,368,633, and 7,439,420 (plant amino acidbiosynthetic enzymes); U.S. Pat. No. 6,459,019 and U.S. application Ser.No. 09/381,485 (increased lysine and threonine); U.S. Pat. No. 6,441,274(plant tryptophan synthase beta subunit); U.S. Pat. No. 6,346,403(methionine metabolic enzymes); U.S. Pat. No. 5,939,599 (high sulfur);U.S. Pat. No. 5,912,414 (increased methionine); U.S. Pat. No. 5,633,436(increasing sulfur amino acid content); U.S. Pat. No. 5,559,223(synthetic storage proteins with defined structure containingprogrammable levels of essential amino acids for improvement of thenutritional value of plants); U.S. Pat. No. 6,194,638 (hemicellulose);U.S. Pat. No. 7,098,381 (UDPGdH); U.S. Pat. No. 6,194,638 (RGP); U.S.Pat. Nos. 6,399,859, 6,930,225, 7,179,955, 6,803,498, 5,850,016, and7,053,282 (alteration of amino acid compositions in seeds); WO 99/29882(methods for altering amino acid content of proteins); U.S. applicationSer. No. 09/297,418 (proteins with enhanced levels of essential aminoacids); WO 98/45458 (engineered seed protein having higher percentage ofessential amino acids); WO 01/79516; and U.S. Pat. Nos. 6,803,498,6,930,225, 7,307,149, 7,524,933, 7,579,443, 7,838,632, 7,851,597, and7,982,009 (maize cellulose synthases).

4. Genes that Control Male Sterility:

There are several methods of conferring genetic male sterilityavailable, such as multiple mutant genes at separate locations withinthe genome that confer male sterility, as disclosed in U.S. Pat. Nos.4,654,465 and 4,727,219 to Brar, et al., and chromosomal translocationsas described by Patterson in U.S. Pat. Nos. 3,861,709 and 3,710,511. Inaddition to these methods, Albertsen, et al., U.S. Pat. No. 5,432,068,describes a system of nuclear male sterility which includes: identifyinga gene which is critical to male fertility; silencing this native genewhich is critical to male fertility; removing the native promoter fromthe essential male fertility gene and replacing it with an induciblepromoter; inserting this genetically engineered gene back into theplant; and thus creating a plant that is male sterile because theinducible promoter is not “on” resulting in the male fertility gene notbeing transcribed. Fertility is restored by inducing, or turning “on,”the promoter, which in turn allows the gene that confers male fertilityto be transcribed.

A. Introduction of a deacetylase gene under the control of atapetum-specific promoter and with the application of the chemicalN-Ac-PPT. See, U.S. Pat. No. 6,384,304.

B. Introduction of various stamen-specific promoters. See, U.S. Pat.Nos. 5,639,948 and 5,589,610.

C. Introduction of the barnase and the barstar genes. See, Paul, et al.,Plant Mol. Biol., 19:611-622 (1992).

For additional examples of nuclear male and female sterility systems andgenes, see also, U.S. Pat. Nos. 5,859,341, 6,297,426, 5,478,369,5,824,524, 5,850,014, and 6,265,640, all of which are herebyincorporated by reference.

5. Genes that Create a Site for Site Specific DNA Integration:

This includes the introduction of FRT sites that may be used in theFLP/FRT system and/or Lox sites that may be used in the Cre/Loxp system.See, for example, Lyznik, et al., Site-Specific Recombination forGenetic Engineering in Plants, Plant Cell Rep, 21:925-932 (2003) andU.S. Pat. No. 6,187,994, which are hereby incorporated by reference.Other systems that may be used include the Gin recombinase of phage Mu(Maeser, et al. (1991); Vicki Chandler, The Maize Handbook, Ch. 118(Springer-Verlag 1994)); the Pin recombinase of E. coli (Enomoto, et al.(1983)); and the R/RS system of the pSRi plasmid (Araki, et al. (1992)).

6. Genes that Affect Abiotic Stress Resistance:

Genes that affect abiotic stress resistance (including but not limitedto flowering, pod and seed development, enhancement of nitrogenutilization efficiency, altered nitrogen responsiveness, droughtresistance or tolerance, cold resistance or tolerance, and saltresistance or tolerance) and increased yield under stress. For example,see U.S. Pat. No. 6,653,535 where water use efficiency is alteredthrough alteration of malate; U.S. Pat. Nos. 5,892,009, 5,965,705,5,929,305, 5,891,859, 6,417,428, 6,664,446, 6,706,866, 6,717,034,6,801,104, 6,946,586, 7,238,860, 7,635,800, 7,135,616, 7,193,129, and7,601,893; and International Publ. Nos. WO 2001/026459, WO 2001/035725,WO 2001/035727, WO 2001/036444, WO 2001/036597, WO 2001/036598, WO2002/015675, and WO 2002/077185, describing genes, including CBF genesand transcription factors effective in mitigating the negative effectsof freezing, high salinity, and drought on plants, as well as conferringother positive effects on plant phenotype; U.S. Publ. No. 2004/0148654,where abscisic acid is altered in plants resulting in improved plantphenotype, such as increased yield and/or increased tolerance to abioticstress; U.S. Pat. Nos. 6,992,237, 6,429,003, 7,049,115, and 7,262,038,where cytokinin expression is modified resulting in plants withincreased stress tolerance, such as drought tolerance, and/or increasedyield. See also, WO 02/02776, WO 2003/052063, JP 2002281975, U.S. Pat.No. 6,084,153, WO 01/64898, and U.S. Pat. Nos. 6,177,275 and 6,107,547(enhancement of nitrogen utilization and altered nitrogenresponsiveness). For ethylene alteration, see, U.S. Publ. Nos.2004/0128719, 2003/0166197, and U.S. application Ser. No. 09/856,834.For plant transcription factors or transcriptional regulators of abioticstress, see, e.g., U.S. Publ. Nos. 2004/0098764 or 2004/0078852.

Other genes and transcription factors that affect plant growth andagronomic traits, such as yield, flowering, plant growth, and/or plantstructure, can be introduced or introgressed into plants. See forexample, U.S. Pat. Nos. 6,140,085, and 6,265,637 (CO); U.S. Pat. No.6,670,526 (ESD4); U.S. Pat. Nos. 6,573,430 and 7,157,279 (TFL); U.S.Pat. No. 6,713,663 (FT); U.S. Pat. Nos. 6,794,560, 6,307,126 (GAI); U.S.Pat. No. 7,045,682 (VRN1); U.S. Pat. Nos. 6,949,694 and 7,253,274(VRN2); U.S. Pat. No. 6,887,708 (GI); U.S. Pat. No. 7,320,158 (FRI);U.S. Pat. No. 6,307,126 (GAI); U.S. Pat. Nos. 6,762,348 and 7,268,272(D8 and Rht); and U.S. Pat. Nos. 7,345,217, 7,511,190, 7,659,446, and7,825,296 (transcription factors).

Genetic Marker Profile Through SSR and First Generation Progeny

In addition to phenotypic observations, a plant can also be identifiedby its genotype. The genotype of a plant can be characterized through agenetic marker profile which can identify plants of the same variety, ora related variety, or be used to determine or validate a pedigree.Genetic marker profiles can be obtained by techniques such asRestriction Fragment Length Polymorphisms (RFLPs), Randomly AmplifiedPolymorphic DNAs (RAPDs), Arbitrarily Primed Polymerase Chain Reaction(AP-PCR), DNA Amplification Fingerprinting (DAF), Sequence CharacterizedAmplified Regions (SCARs), Amplified Fragment Length Polymorphisms(AFLPs), Simple Sequence Repeats (SSRs) (which are also referred to asMicrosatellites), and Single Nucleotide Polymorphisms (SNPs). Forexample, see, Cregan, et al., “An Integrated Genetic Linkage Map of theSoybean Genome,” Crop Science, 39:1464-1490 (1999) and Berry, et al.,“Assessing Probability of Ancestry Using Simple Sequence RepeatProfiles: Applications to Maize Inbred Lines and Soybean Varieties,”Genetics, 165:331-342 (2003), each of which are incorporated byreference herein in their entirety.

Particular markers used for these purposes are not limited to anyparticular set of markers, but are envisioned to include any type ofmarker and marker profile which provides a means of distinguishingvarieties. One method of comparison is to use only homozygous loci forsoybean cultivar 2163154.

Primers and PCR protocols for assaying these and other markers aredisclosed in the Soybase (sponsored by the USDA Agricultural ResearchService and Iowa State University). In addition to being used foridentification of soybean variety 2163154, and plant parts and plantcells of soybean variety 2163154, the genetic profile may be used toidentify a soybean plant produced through the use of soybean cultivar2163154 or to verify a pedigree for progeny plants produced through theuse of soybean cultivar 2163154. The genetic marker profile is alsouseful in breeding and developing backcross conversions.

One embodiment comprises a soybean plant characterized by molecular andphysiological data obtained from the sample of said variety depositedwith the American Type Culture Collection (ATCC) or with the NationalCollections of Industrial, Food and Marine Bacteria (NCIMB). Furtherprovided by the embodiment(s) is a soybean plant formed by thecombination of the disclosed soybean plant or plant cell with anothersoybean plant or cell and comprising the homozygous alleles of thevariety. “Cell” as used herein includes a plant cell, whether isolated,in tissue culture or incorporated in a plant or plant part.

Means of performing genetic marker profiles using SSR polymorphisms arewell known in the art. SSRs are genetic markers based on polymorphismsin repeated nucleotide sequences, such as microsatellites. A markersystem based on SSRs can be highly informative in linkage analysisrelative to other marker systems in that multiple alleles may be present(“linkage” refers to a phenomenon wherein alleles on the same chromosometend to segregate together more often than expected by chance if theirtransmission was independent). Another advantage of this type of markeris that, through use of flanking primers, detection of SSRs can beachieved, for example, by the polymerase chain reaction (PCR), therebyeliminating the need for labor-intensive Southern hybridization. The PCRdetection is done by use of two oligonucleotide primers flanking thepolymorphic segment of repetitive DNA. Repeated cycles of heatdenaturation of the DNA followed by annealing of the primers to theircomplementary sequences at low temperatures, and extension of theannealed primers with DNA polymerase, comprise the major part of themethodology.

Following amplification, markers can be scored by electrophoresis of theamplification products. Scoring of marker genotype is based on the sizeof the amplified fragment, which may be measured by the number of basepairs of the fragment. While variation in the primer used or inlaboratory procedures can affect the reported fragment size, relativevalues should remain constant regardless of the specific primer orlaboratory used. When comparing varieties, it is preferable if all SSRprofiles are performed in the same lab.

Primers used are publicly available and may be found in the Soybase orCregan supra. See also, U.S. application Ser. No. 09/581,970 (NucleotidePolymorphisms in Soybean); U.S. Pat. No. 6,162,967 (Positional Cloningof Soybean Cyst Nematode Resistance Genes); and U.S. Pat. No. 7,288,386(Soybean Sudden Death Syndrome Resistant Soybeans and Methods ofBreeding and Identifying Resistant Plants), the disclosure of which areincorporated herein by reference.

The SSR profile of soybean plant 2163154 can be used to identify plantscomprising soybean cultivar 2163154 as a parent, since such plants willcomprise the same homozygous alleles as soybean cultivar 2163154.Because the soybean variety is essentially homozygous at all relevantloci, most loci should have only one type of allele present. Incontrast, a genetic marker profile of an F₁ progeny should be the sum ofthose parents, e.g., if one parent was homozygous for allele x at aparticular locus, and the other parent homozygous for allele y at thatlocus, then the F₁ progeny will be xy (heterozygous) at that locus.Subsequent generations of progeny produced by selection and breeding areexpected to be of genotype x (homozygous), y (homozygous), or xy(heterozygous) for that locus position. When the F₁ plant is selfed orsibbed for successive filial generations, the locus should be either xor y for that position.

In addition, plants and plant parts substantially benefiting from theuse of soybean cultivar 2163154 in their development, such as soybeancultivar 2163154 comprising a backcross conversion, transgene, orgenetic sterility factor, may be identified by having a molecular markerprofile with a high percent identity to soybean cultivar 2163154. Such apercent identity might be 95%, 96%, 97%, 98%, 99%, 99.5%, or 99.9%identical to soybean cultivar 2163154. Percent identity refers to thecomparison of the homozygous alleles of two soybean varieties. Percentidentity or percent similarity is determined by comparing astatistically significant number of the homozygous alleles of twodeveloped varieties. For example, a percent identity of 90% betweensoybean variety 1 and soybean variety 2 means that the two varietieshave the same allele at 90% of their loci.

The SSR profile of soybean cultivar 2163154 can also be used to identifyessentially derived varieties and other progeny varieties developed fromthe use of soybean cultivar 2163154, as well as cells and other plantparts thereof. Such plants may be developed using the markers, forexample, identified in U.S. Pat. Nos. 6,162,967, and 7,288,386. Progenyplants and plant parts produced using soybean cultivar 2163154 may beidentified by having a molecular marker profile of at least 25%, 30%,35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 76%, 77%, 78%, 79%, 80%,81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98%, 99%, or 99.5% genetic contribution from soybeanvariety, as measured by either percent identity or percent similarity.Such progeny may be further characterized as being within a pedigreedistance of soybean cultivar 2163154, such as within 1, 2, 3, 4, or 5 orless cross-pollinations to a soybean plant other than soybean cultivar2163154 or a plant that has soybean cultivar 2163154 as a progenitor.Unique molecular profiles may be identified with other molecular toolssuch as SNPs and RFLPs.

While determining the SSR genetic marker profile of the plants describedsupra, several unique SSR profiles may also be identified which did notappear in either parent of such plant. Such unique SSR profiles mayarise during the breeding process from recombination or mutation. Acombination of several unique alleles provides a means of identifying aplant variety, an F₁ progeny produced from such variety, and progenyproduced from such variety.

Tissue Culture

Further reproduction of the variety can occur by tissue culture andregeneration. Tissue culture of various tissues of soybeans andregeneration of plants therefrom is well-known and widely published. Forexample, reference may be had to Komatsuda, T., et al., Crop Sci.,31:333-337 (1991); Stephens, P. A., et al., Theor. Appl. Genet.,82:633-635 (1991); Komatsuda, T., et al., Plant Cell, Tissue and OrganCulture, 28:103-113 (1992); Dhir, S., et al., Plant Cell Reports,11:285-289 (1992); Pandey, P., et al., Japan J. Breed., 42:1-5 (1992);and Shetty, K., et al., Plant Science, 81:245-251 (1992); as well asU.S. Pat. Nos. 5,024,944 and 5,008,200. Thus, another aspect orembodiment is to provide cells which upon growth and differentiationproduce soybean plants having the physiological and morphologicalcharacteristics of soybean cultivar 2163154.

Regeneration refers to the development of a plant from tissue culture.The term “tissue culture” indicates a composition comprising isolatedcells of the same or a different type or a collection of such cellsorganized into parts of a plant. Exemplary types of tissue cultures areprotoplasts, calli, plant clumps, and plant cells that can generatetissue culture that are intact in plants or parts of plants, such asembryos, pollen, flowers, seeds, pods, petioles, leaves, stems, roots,root tips, anthers, pistils, and the like. Means for preparing andmaintaining plant tissue culture are well known in the art. By way ofexample, a tissue culture comprising organs has been used to produceregenerated plants. U.S. Pat. Nos. 5,959,185, 5,973,234, and 5,977,445describe certain techniques, the disclosures of which are incorporatedherein by reference.

Industrial Uses

The seed of soybean cultivar 2163154, the plant produced from the seed,the hybrid soybean plant produced from the crossing of the variety withany other soybean plant, hybrid seed, and various parts of the hybridsoybean plant can be utilized for human food, livestock feed, and as araw material in industry. The soybean seeds produced by soybean cultivar2163154 can be crushed, or a component of the soybean seeds can beextracted, in order to comprise a commodity plant product, such asprotein concentrate, protein isolate, soybean hulls, meal, flour, or oilfor a food or feed product.

The soybean is the world's leading source of vegetable oil and proteinmeal. The oil extracted from soybeans is used for cooking oil,margarine, and salad dressings. Soybean oil is composed of saturated,monounsaturated, and polyunsaturated fatty acids. It has a typicalcomposition of 11% palmitic, 4% stearic, 25% oleic, 50% linoleic, and 9%linolenic fatty acid content (“Economic Implications of Modified SoybeanTraits Summary Report,” Iowa Soybean Promotion Board and AmericanSoybean Association Special Report 92S (May 1990)). Changes in fattyacid composition for improved oxidative stability and nutrition areconstantly sought after. Industrial uses of soybean oil, which issubjected to further processing, include ingredients for paints,plastics, fibers, detergents, cosmetics, lubricants, and biodiesel fuel.Soybean oil may be split, inter-esterified, sulfurized, epoxidized,polymerized, ethoxylated, or cleaved. Designing and producing soybeanoil derivatives with improved functionality and improved oliochemistryis a rapidly growing field. The typical mixture of triglycerides isusually split and separated into pure fatty acids, which are thencombined with petroleum-derived alcohols or acids, nitrogen, sulfonates,chlorine, or with fatty alcohols derived from fats and oils to producethe desired type of oil or fat.

Soybean cultivar 2163154 can be used to produce soybean oil. To producesoybean oil, the soybeans harvested from soybean cultivar 2163154 arecracked, adjusted for moisture content, rolled into flakes and the oilis solvent-extracted from the flakes with commercial hexane. The oil isthen refined, blended for different applications, and sometimeshydrogenated. Soybean oils, both liquid and partially hydrogenated, areused domestically and exported, sold as “vegetable oil” or are used in awide variety of processed foods.

Soybeans are also used as a food source for both animals and humans.Soybeans are widely used as a source of protein for poultry, swine, andcattle feed. During processing of whole soybeans, the fibrous hull isremoved and the oil is extracted. The remaining soybean meal is acombination of carbohydrates and approximately 50% protein.

Soybean cultivar 2163154 can be used to produce meal. After oil isextracted from whole soybeans harvested from soybean cultivar 2163154,the remaining material or “meal” is “toasted” (a misnomer because theheat treatment is with moist steam) and ground in a hammer mill. Soybeanmeal is an essential element of the American production method ofgrowing farm animals, such as poultry and swine, on an industrial scalethat began in the 1930s; and more recently the aquaculture of catfish.Ninety-eight percent of the U.S. soybean crop is used for livestockfeed. Soybean meal is also used in lower end dog foods. Soybean mealproduced from soybean cultivar 2163154 can also be used to producesoybean protein concentrate and soybean protein isolate.

In addition to soybean meal, soybean cultivar 2163154 can be used toproduce soy flour. Soy flour refers to defatted soybeans where specialcare was taken during desolventizing (not toasted) to minimizedenaturation of the protein and to retain a high Nitrogen SolubilityIndex (NSI) in making the flour. Soy flour is the starting material forproduction of soy concentrate and soy protein isolate. Defatted soyflour is obtained from solvent extracted flakes, and contains less than1% oil. Full-fat soy flour is made from unextracted, dehulled beans, andcontains about 18% to 20% oil. Due to its high oil content, aspecialized Alpine Fine Impact Mill must be used for grinding ratherthan the more common hammer mill. Low-fat soy flour is made by addingback some oil to defatted soy flour. The lipid content varies accordingto specifications, usually between 4.5% and 9%. High-fat soy flour canalso be produced by adding back soybean oil to defatted flour at thelevel of 15%. Lecithinated soy flour is made by adding soybean lecithinto defatted, low-fat or high-fat soy flours to increase theirdispersibility and impart emulsifying properties. The lecithin contentvaries up to 15%.

For human consumption, soybean cultivar 2163154 can be used to produceedible protein ingredients which offer a healthier, less expensivereplacement for animal protein in meats, as well as in dairy-typeproducts. The soybeans produced by soybean cultivar 2163154 can beprocessed to produce a texture and appearance similar to many otherfoods. For example, soybeans are the primary ingredient in many dairyproduct substitutes (e.g., soy milk, margarine, soy ice cream, soyyogurt, soy cheese, and soy cream cheese) and meat substitutes (e.g.,veggie burgers). These substitutes are readily available in mostsupermarkets. Although soy milk does not naturally contain significantamounts of digestible calcium (the high calcium content of soybeans isbound to the insoluble constituents and remains in the soy pulp), manymanufacturers of soy milk sell calcium-enriched products as well. Soy isalso used in tempe, where the beans (sometimes mixed with grain) arefermented into a solid cake.

Additionally, soybean cultivar 2163154 can be used to produce varioustypes of “fillers” in meat and poultry products. Food service, retail,and institutional (primarily school lunch and correctional) facilitiesregularly use such “extended” products, that is, products which containsoy fillers. Extension may result in diminished flavor, but fat andcholesterol are reduced by adding soy fillers to certain products.Vitamin and mineral fortification can be used to make soy productsnutritionally equivalent to animal protein; the protein quality isalready roughly equivalent.

While a number of exemplary aspects and embodiments have been discussedabove, those of skill in the art will recognize certain modifications,permutations, additions and sub-combinations thereof. It is thereforeintended that the following appended claims and claims hereafterintroduced are interpreted to include all such modifications,permutations, additions, and sub-combinations as are within their truespirit and scope.

One embodiment may be embodied in other specific forms without departingfrom its spirit or essential characteristics. The described embodimentsare to be considered in all respects only as illustrative and notrestrictive. All changes which come within the meaning and range ofequivalency of the claims are to be embraced within their scope.

Various embodiments, include components, methods, processes, systemsand/or apparatus substantially as depicted and described herein,including various embodiments, sub-combinations, and subsets thereof.Those of skill in the art will understand how to make and use anembodiment(s) after understanding the present disclosure.

The foregoing discussion of the embodiments has been presented forpurposes of illustration and description. The foregoing is not intendedto limit the embodiments to the form or forms disclosed herein. In theforegoing Detailed Description for example, various features of theembodiments are grouped together in one or more embodiments for thepurpose of streamlining the disclosure. This method of disclosure is notto be interpreted as reflecting an intention that the embodiment(s)requires more features than are expressly recited in each claim. Rather,as the following claims reflect, inventive aspects lie in less than allfeatures of a single foregoing disclosed embodiment. Thus, the followingclaims are hereby incorporated into this Detailed Description.

Moreover, though the description of the embodiments has includeddescription of one or more embodiments and certain variations andmodifications, other variations and modifications are within the scopeof the embodiments (e.g., as may be within the skill and knowledge ofthose in the art, after understanding the present disclosure). It isintended to obtain rights which include alternative embodiments to theextent permitted, including alternate, interchangeable and/or equivalentstructures, functions, ranges or acts to those claimed, whether or notsuch alternate, interchangeable and/or equivalent structures, functions,ranges or acts are disclosed herein, and without intending to publiclydedicate any patentable subject matter.

The use of the terms “a,” “an,” and “the,” and similar referents in thecontext of describing the embodiments (especially in the context of thefollowing claims) are to be construed to cover both the singular and theplural, unless otherwise indicated herein or clearly contradicted bycontext. The terms “comprising,” “having,” “including,” and “containing”are to be construed as open-ended terms (i.e., meaning “including, butnot limited to,”) unless otherwise noted. Recitation of ranges of valuesherein are merely intended to serve as a shorthand method of referringindividually to each separate value falling within the range, unlessotherwise indicated herein, and each separate value is incorporated intothe specification as if it were individually recited herein. Forexample, if the range 10-15 is disclosed, then 11, 12, 13, and 14 arealso disclosed. All methods described herein can be performed in anysuitable order unless otherwise indicated herein or otherwise clearlycontradicted by context. The use of any and all examples, or exemplarylanguage (e.g., “such as”) provided herein, is intended merely to betterilluminate the embodiments and does not pose a limitation on the scopeof the embodiments unless otherwise claimed.

DEPOSIT INFORMATION

A deposit of the SGI Genetics, Inc. proprietary soybean cultivar 2163154disclosed above and recited in the appended claims is maintained by SGIGenetics, Inc. A deposit will be made with the National Collections ofIndustrial, Food and Marine Bacteria (NCIMB), Ferguson Building,Craibstone Estate, Bucksburn, Aberdeen, AB21 9YA, Scotland, UnitedKingdom. Access to this deposit will be available during the pendency ofthis application to persons determined by the Commissioner of Patentsand Trademarks to be entitled thereto under 37 C.F.R. 1.14 and 35 U.S.C.§ 122. Upon allowance of any claims in this application, allrestrictions on the availability to the public of the variety will beirrevocably removed by affording access to a deposit of at least 2,500seeds of the same variety with NCIMB. The deposit will be maintained inthe depository for a period of 30 years, or 5 years after the lastrequest, or for the effective life of the patent, whichever is longer,and will be replaced if necessary, during that period.

What is claimed is:
 1. A plant or a seed of soybean cultivar 2163154,wherein a representative sample of seed of said cultivar is depositedunder NCIMB No. ______.
 2. A soybean plant, or a part thereof, producedby growing a seed of soybean cultivar 2163154, wherein a representativesample of seed of said cultivar is deposited under NCIMB No. _______. 3.A cell of the plant or seed of claim
 1. 4. A tissue culture ofprotoplasts or regenerable cells from the cell of claim
 3. 5. A soybeanplant regenerated from tissue culture of claim
 4. 6. A method forproducing a soybean seed, comprising crossing two soybean plants andharvesting the resultant soybean seed, wherein at least one soybeanplant is the soybean plant of claim
 1. 7. An F₁ soybean seed produced bythe method of claim
 6. 8. A soybean plant, or a part thereof, producedby growing the seed of claim
 7. 9. A method of producing a plant ofsoybean cultivar 2163154 comprising an added desired trait, the methodcomprising the step of introducing at least one gene or locus conferringthe desired trait into the plant of claim
 1. 10. A plant produced by themethod of claim 9, wherein the plant comprises the desired trait andessentially all of the physiological and morphological characteristicsof soybean cultivar
 2163154. 11. A method of introducing a desired traitinto soybean cultivar 2163154, wherein the method comprises: (a)crossing a 2163154 plant, wherein a sample of seed is deposited underNCIMB No. ______, with a plant of another soybean cultivar having adesired trait to produce progeny plants, wherein the desired trait ischosen from male sterility, herbicide resistance, insect resistance,modified fatty acid metabolism, modified carbohydrate metabolism,modified seed yield, modified seed oil, modified seed protein, modifiedlodging resistance, modified shattering, modified iron-deficiencychlorosis and resistance to herbicides, insects, bacterial disease,fungal disease or viral disease; (b) selecting one or more progenyplants that have the desired trait to produce selected progeny plants;(c) crossing the selected progeny plants with the 2163154 plant toproduce backcross progeny plants; (d) selecting for backcross progenyplants that have the desired trait; and (e) repeating steps (c) and (d)a sufficient number of times in succession to produce selected second orhigher backcross progeny plants that comprise the desired trait andessentially all of the physiological and morphological characteristicsof soybean cultivar
 2163154. 12. A soybean plant produced by the methodof claim 11 wherein the plant has the desired trait.
 13. A method ofproducing a soybean plant with modified fatty acid metabolism ormodified carbohydrate metabolism, wherein the method comprisesintroducing a gene encoding a protein chosen from phytase,fructosyltransferase, levansucrase, α-amylase, invertase and starchbranching enzyme or encoding an antisense polynucleotide effective forinhibition of expression of stearyl-ACP desaturase into the soybeanplant of claim
 1. 14. A soybean plant having modified fatty acidmetabolism or modified carbohydrate metabolism produced by the method ofclaim 13, and wherein said plant comprises essentially all of thephysiological and morphological characteristics of soybean cultivar2163154 listed in Table
 1. 15. A method of producing an herbicideresistant soybean plant, wherein the method comprises introducing a geneconferring herbicide resistance into the plant of claim
 1. 16. Anherbicide resistant soybean plant produced by the method of claim 15,wherein the gene confers resistance to an herbicide selected from thegroup consisting of glyphosate, sulfonylurea, imidazolinone, dicamba,glufosinate, phenoxy proprionic acid, L-phosphinothricin, cyclohexone,cyclohexanedione, triazine, PPO-herbicides, bromoxynil, andbenzonitrile, and wherein said plant comprises essentially all of thephysiological and morphological characteristics of soybean cultivar2163154 listed in Table
 1. 17. A method of producing a pest or insectresistant soybean plant, wherein the method comprises introducing a geneconferring pest or insect resistance into the soybean plant of claim 1,and wherein said plant comprises essentially all of the physiologicaland morphological characteristics of soybean cultivar 2163154 listed inTable
 1. 18. A pest or insect resistant soybean plant produced by themethod of claim 17, and wherein said plant comprises essentially all ofthe physiological and morphological characteristics of soybean cultivar2163154 listed in Table
 1. 19. The soybean plant of claim 18, whereinthe gene encodes a Bacillus thuringiensis (Bt) endotoxin, and whereinsaid plant comprises essentially all of the physiological andmorphological characteristics of soybean cultivar 2163154 listed inTable
 1. 20. A method of producing a disease resistant soybean plant,wherein the method comprises introducing a gene which confers diseaseresistance into the soybean plant of claim 1, and wherein said plantcomprises essentially all of the physiological and morphologicalcharacteristics of soybean cultivar 2163154 listed in Table
 1. 21. Adisease resistant soybean plant produced by the method of claim 20, andwherein said plant comprises essentially all of the physiological andmorphological characteristics of soybean cultivar 2163154 listed inTable
 1. 22. A method of producing a commodity plant product, comprisingobtaining the plant of claim 1, or a part thereof, and producing thecommodity plant product from the plant or part thereof, wherein thecommodity plant product is selected from the group consisting of proteinconcentrate, protein isolate, soybean hulls, meal, flour and oil.
 23. Amethod for developing a soybean plant, comprising applying plantbreeding techniques to the plant of claim 1, or plant part thereof,comprising crossing, recurrent selection, mutation breeding, whereinsaid mutation breeding selects for a mutation that is spontaneous orartificially induced, backcrossing, pedigree breeding, marker enhancedselection, haploid/double haploid production, or transformation, whereinapplication of said techniques results in development of a new soybeanplant.
 24. A method of introducing a mutation into the genome of soybeancultivar 2163154, said method comprising mutagenesis of the plant ofclaim 1, or plant part thereof, wherein said mutagenesis is selectedfrom the group consisting of temperature, long-term seed storage, tissueculture conditions, ionizing radiation, chemical mutagens, or targetinginduced local lesions in genomes, and wherein the resulting plantcomprises at least one genome mutation.
 25. A method of editing thegenome of soybean cultivar 2163154, said method comprising editing thegenome of the plant, or plant part thereof, of claim 2, wherein saidmethod is selected from the group comprising zinc finger nucleases,transcription activator-like effector nucleases (TALENs), engineeredhoming endonucleases/meganucleases, and the clustered regularlyinterspaced short palindromic repeat (CRISPR)-associated protein9 (Cas9)system.
 26. A soybean plant produced by the method of claim
 25. 27. Amethod of producing a commodity plant product, comprising obtaining theplant of claim 1, or a part thereof, and producing the commodity plantproduct from the plant or part thereof, wherein the commodity plantproduct is selected from the group consisting of protein concentrate,protein isolate, soybean hulls, meal, flour and oil.
 28. A soybeancommodity plant product produced from the plant or seed of claim 1,wherein the commodity plant product comprises at least one cell ofsoybean cultivar 2163154.